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/ß/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.

Investigating and combatting the key drivers of viral zoonoses in Africa: an analysis of eight epidemics / Investigar e combater os principais promotores de zoonoses virais na África: uma análise de oito epidemias

Investigating the interplay of factors that result in a viral zoonotic outbreak is difficult, though it is increasingly important. As anthropogenic influences shift the delicate balance of ecosystems, new zoonoses emerge in humans. Sub-Saharan Africa is a notable hotspot for zoonotic disease due to abundant competent mammalian reservoir hosts. Furthermore, poverty, corruption, and an overreliance on natural resources play considerable roles in depleting biological resources, exacerbating the population's susceptibility. Unsurprisingly, viral zoonoses have emerged in Africa, including HIV/AIDS, Ebola, Avian influenza, Lassa fever, Zika, and Monkeypox. These diseases are among the principal causes of death in endemic areas. Though typically distinct in their manifestations, viral zoonoses are connected by underlying, definitive factors. This review summarises vital findings on viral zoonoses in Africa using nine notable case studies as a benchmark for future studies. We discuss the importance of ecological recuperation and protection as a central strategy to control zoonotic diseases. Emphasis was made on moderating key drivers of zoonotic diseases to forestall future pandemics. This is in conjunction with attempts to redirect efforts from reactive to pre-emptive through a multidisciplinary "one health" approach.

Investigar a interação de fatores que resultam em um surto zoonótico viral é difícil, embora seja cada vez mais relevante. À medida que as influências antropogênicas mudam o delicado equilíbrio dos ecossistemas, novas zoonoses surgem em humanos. A África Subsaariana é um ponto crítico notável para doenças zoonóticas devido a abundantes reservatórios mamíferos competentes. Além disso, a pobreza, a corrupção e o excesso de confiança nos recursos naturais desempenham papéis consideráveis no esgotamento dos recursos biológicos, exacerbando a suscetibilidade da população. Sem surpresa, zoonoses virais surgiram na África, incluindo HIV/AIDS, Ebola, gripe aviária, febre de lassa, zika e varíola dos macacos. Essas doenças estão entre as principais causas de morte em áreas endêmicas. Apesar de serem tipicamente distintas em suas manifestações, as zoonoses virais estão conectadas por fatores subjacentes e definitivos. Esta revisão resume descobertas vitais sobre zoonoses virais na África usando nove estudos de caso notáveis como referência para estudos futuros. Discutimos a importância da recuperação e proteção ecológica como estratégia central para o controle de doenças zoonóticas. Foi dada ênfase à moderação dos principais impulsionadores de doenças zoonóticas para prevenir futuras pandemias. Isso ocorre em conjunto com tentativas de redirecionar os esforços de reativos para preventivos por meio de uma abordagem multidisciplinar de "uma só saúde".

West Nile virus in Mexico: Why vectors matter for explaining the current absence of epidemics.

Since 2002, West Nile Virus (WNV) has been reported in 18 states in Mexico, either by PCR or serological testing. However, it is believed that the virus is present in more states. Only four states (out of 32) have reported confirmed human cases, and one state has serological evidence. In the country, WNV is present in mainly horses and birds, but its presence extends to crocodiles, felines, canines, swines, donkeys, caprines, antilopes, cattle, bats, and camelids. Positive mosquito species include Aedes and Culex spp. Different hypotheses have been proposed to explain the absence of WNV epidemics in Latin America. Since some regions of Mexico and the United States share ecological and climatic conditions, these hypotheses may not be sufficient to account for the absence of WNV outbreaks or epidemics. This paper discusses the proposed ideas and attempts to contextualize them for Mexico, particularly for the U.S.-Mexico border, where WNV infections have been reported in humans, horses, and mosquitoes. We propose that integration of urban ecology and entomology knowledge is needed to better understand the absence of WN cases in Mexico.

Vector Competence of Culex quinquefasciatus from Brazil for West Nile Virus.

West Nile virus is characterized as a neurotropic pathogen, which can cause West Nile fever and is transmitted by mosquitoes of the genus Culex. In 2018, the Instituto Evandro Chagas performed the first isolation of a WNV strain in Brazil from a horse brain sample. The present study aimed to evaluate the susceptibility of orally infected Cx. quinquefasciatus from the Amazon region of Brazil to become infected and transmit the WNV strain isolated in 2018. Oral infection was performed with blood meal artificially infected with WNV, followed by analysis of infection, dissemination, and transmission rates, as well as viral titers of body, head, and saliva samples. At the 21st dpi, the infection rate was 100%, the dissemination rate was 80%, and the transmission rate was 77%. These results indicate that Cx. quinquefasciatus is susceptible to oral infection by the Brazilian strain of WNV and may act as a possible vector of the virus since it was detected in saliva from the 21st dpi.

First Isolation and Genome Sequence Analysis of West Nile Virus in Mosquitoes in Brazil.

West Nile virus is a flavivirus transmitted by mosquitoes, mainly of the genus Culex. In Brazil, serological studies have already indicated the circulation of the virus since 2003, with the first human case detected in 2014. The objective of the present paper is to report the first isolation of WNV in a Culex (Melanoconion) mosquito. Arthropods were collected by protected human attraction and CDC light bait, and taxonomically identified and analyzed by viral isolation, complement fixation and genomic sequencing tests. WNV was isolated from samples of Culex (Melanoconion) mosquitoes, and the sequencing analysis demonstrated that the isolated strain belonged to lineage 1a. The finding of the present study presents the first evidence of the isolation and genome sequencing of WNV in arthropods in Brazil.

Arbovirus surveillance on the Mexico-USA border: West Nile virus identification in various species of field mosquitoes.

West Nile virus (WNV) has been documented in human and/or mosquito samples near the border with Mexico in El Paso, Texas, and Doña Ana County, New Mexico. However, on the Mexican side of the border, particularly in the State of Chihuahua, no such cases of WNV-infected mosquitoes have been documented. We tested 367 mosquitoes of four species (Culex quinquefasciatus, Cx. tarsalis, Aedes aegypti, and Aedes (Ochlerotatus) epactius) and found a high rate of WNV-positivity, including the first record of Ae. (Ochlerotatus) epactius infection with WNV. These results call for intensifying WNV surveillance efforts on the border between the United States and Mexico, with particular emphasis on vector control and monitoring of the species included in this study.

Arboviruses in Mammals in the Neotropics: A Systematic Review to Strengthen Epidemiological Monitoring Strategies and Conservation Medicine.

Arthropod-borne viruses (arboviruses) are a diverse group of ribonucleic acid (RNA) viruses, with the exception of African swine fever virus, that are transmitted by hematophagous arthropods to a vertebrate host. They are the important cause of many diseases due to their ability to spread in different environments and their diversity of vectors. Currently, there is no information on the geographical distribution of the diseases because the routes of transmission and the mammals (wild or domestic) that act as potential hosts are poorly documented or unknown. We conducted a systematic review from 1967 to 2021 to identify the diversity of arboviruses, the areas, and taxonomic groups that have been monitored, the prevalence of positive records, and the associated risk factors. We identified forty-three arboviruses in nine mammalian orders distributed in eleven countries. In Brazil, the order primates harbor the highest number of arbovirus records. The three most recorded arboviruses were Venezuelan equine encephalitis, Saint Louis encephalitis and West Nile virus. Serum is the most used sample to obtain arbovirus records. Deforestation is identified as the main risk factor for arbovirus transmission between different species and environments (an odds ratio of 1.46 with a 95% confidence interval: 1.34-1.59). The results show an increase in the sampling effort over the years in the neotropical region. Despite the importance of arboviruses for public health, little is known about the interaction of arboviruses, their hosts, and vectors, as some countries and mammalian orders have not yet been monitored. Long-term and constant monitoring allows focusing research on the analysis of the interrelationships and characteristics of each component animal, human, and their environment to understand the dynamics of the diseases and guide epidemiological surveillance and vector control programs. The biodiversity of the Neotropics should be considered to support epidemiological monitoring strategies.

Serological Determination of West Nile Virus in Domestic Birds from Rapa Nui, Chile.

Background: Flaviviruses are agents with high zoonotic potential of importance to human health. They are transmitted by mosquitoes of the Culicidae family, and birds act as host-amplifiers. Birds, mammals, and humans are susceptible hosts to infection. Methods: In this study, West Nile virus (WNV), flavivirus, infection was studied in 37 serum samples from 22 hens on Easter Island, Chile. Results: WNV was detected by ELISA (ID Screen® West Nile Competition Multi-Species). We report absence of antibodies to WNV, and to related viruses of the Japanese Encephalitis Virus serocomplex, and, therefore, absence of infection across the sample. Conclusion: This is the first evaluation of its type carried out in Chile, and represents a positive result for public health at Easter Island.

Blood feeding patterns of the Culex pipiens complex in equestrian land uses and their implications for arboviral encephalitis risk in temperate Argentina.

Blood feeding patterns of mosquitoes are a key component in the dynamics of arboviral encephalitides transmission. In temperate Argentina, the members of the Culex pipiens complex include Cx. pipiens molestus, Cx. quinquefasciatus and their hybrids. To characterize their blood feeding patterns, adult resting mosquitoes were collected monthly during the warm season in urban and rural equestrian fields. The availability of birds and domestic mammals per site was characterized. The blood source and the complex member were successfully identified for 89 specimens using PCR. Blood of 19 vertebrate species was isolated including four mammals (most common feeds from dog, Canus lupus 19% of the blood meals; and horse, Equus caballus 18%) and 15 birds (picazuro pigeon, Patagioenas picazuro 11%; eared dove, Zenaida auriculata 10%; chicken, Gallus gallus 9%). The Forage Ratio (FR), calculated as the proportion of feeds taken from a given host species with respect to that host availability in the environment, suggested preference for dog by all members of the complex (FR ≥4.5). On the contrary, FR values suggested avoidance for horse by Cx. quinquefasciatus and the hybrid (FR ≤0.8), and a use proportional to its abundance by Cx. pipiens molestus (FR = 1.1-1.2 in urban and rural sites, respectively). FR values suggesting preference were obtained for avian species of the orders Passeriformes (7 species in total) and Columbiformes (5) by all members of the complex (FR ≥ 3.3), whereas values for monk parakeet (Myiopsitta monachus, Psiitaciformes) suggested avoidance by Cx. quinquefasciatus in urban sites (FR = 0.4) and by Cx. pipiens molestus in rural sites (FR = 0.3) but not in urban sites (FR = 1.4). A mammal-bird index (MBI, from -1 all avian to +1 all mammalian blood meals) was calculated for each member of the complex and urbanization category. Values were negative for Cx. quinquefasciatus (MBIurban  = -0.60, MBIrural  = -0.33) and positive for Cx. pipiens molestus (MBIurban  = 0.20, MBIrural  = 0.60), indicating a higher proportion of feeds taken on birds and mammals, respectively, regardless of the urbanization category. In temperate Argentina, the members of the Cx. pipiens complex fed both on horses and on birds, thus representing a real risk of transmission of arboviral encephalitides from avian enzootic cycles to horse epizootics.

Co-infection of Peruvian horse sickness virus and West Nile virus associated with neurological diseases in horses from Brazil.

In 2018, during the surveillance for West Nile virus (WNV) in horses with neurological clinical signs in the state of Espírito Santo (Brazil), 19 animals were investigated, and 52 biological samples were collected for WNV diagnostic. One brain sample was positive for WNV by RT-qPCR and the virus was isolated in C6/36 cell culture and sequenced. We obtained a nearly complete genome of WNV co-infected with Peruvian horse sickness virus (PHSV) in the cell culture. After confirmation of PHSV by next-generation sequencing, a new PHSV RT-qPCR protocol was developed, which was used to detect another horse positive only for PHSV. This assay provides a simple and direct method for easy identification of PHSV from biological samples from horses and may become a useful tool in the epidemiological surveillance of this virus. It is the first case of PHSV in Brazil, and only the third country overall to report, 23 years after the first confirmed notification in Peru. Moreover, it is the first reported co-infection of PHSV and WNV in a horse with neurological signs, confirmed by RT-qPCR.

Serological Evidence of Exposure to Saint Louis Encephalitis and West Nile Viruses in Horses of Rio de Janeiro, Brazil.

Infections with arboviruses are reported worldwide. Saint Louis encephalitis (SLEV) and West Nile (WNV) viruses are closely related flaviviruses affecting humans and animals. SLEV has been sporadically detected in humans, and corresponding antibodies have been frequently detected in horses throughout Brazil. WNV was first reported in western Brazil over a decade ago, has been associated with neurological disorders in humans and equines and its prevalence is increasing nationwide. Herein, we investigated by molecular and serological methods the presence of SLEV and WNV in equines from Rio de Janeiro. A total of 435 serum samples were collected from healthy horses and tested for specific neutralizing antibodies by plaque reduction neutralization test (PRNT90). Additionally, samples (serum, cerebrospinal fluid, central nervous system tissue) from 72 horses, including horses with neurological disorders resulting in a fatal outcome or horses which had contact with them, were tested by real-time reverse transcription-polymerase chain reaction (RT-qPCR) for both viruses. Adopting the criterion of four-fold antibody titer difference, 165 horses (38%) presented neutralizing antibodies for flaviviruses, 89 (20.4%) for SLEV and five (1.1%) for WNV. No evidence of SLEV and WNV infection was detected by RT-qPCR and, thus, such infection could not be confirmed in the additional samples. Our findings indicate horses of Rio de Janeiro were exposed to SLEV and WNV, contributing to the current knowledge on the distribution of these viruses in Brazil.

West Nile Virus Surveillance in Sentinel Chickens and Mosquitoes in Panama City Beach, Florida, from 2014 To 2020.

Over 20 years since its introduction, the West Nile virus (WNV) continues to be the leading cause of arboviral disease in the USA. In Panama City Beach (Bay County, FL), WNV transmission is monitored using sentinel chickens and testing mosquito pools for presence of viral RNA. In the current work, we monitored WNV transmission from 2014 to 2020 through weekly serology sampling of sentinel chickens; mosquito populations through biweekly mosquito collections by suction traps (1 m and 9 m) and weekly gravid trap collections; and mosquito infection rates using a reverse transcriptase-polymerase chain reaction (RT-PCR) assay. Samples were sent to the Bureau of Public Health Laboratories (Tampa, FL) for testing presence/absence of WNV via RT-PCR assay. Our results indicated that canopy surveillance could augment ground collections, providing greater proportions of Culex mosquitoes with less bycatch compared with ground collections. Serology indicated 94 seroconversions to WNV in the study area from 2014 to 2020. The most active year was 2016, which accounted for 32% (n = 30) of all seroconversions reported during the study period. We detected 20 WNV-positive mosquito pools from Culex quinquefasciatus during 2014-17; mosquito infection rates ranged from 2.02 to 23.81 per thousand (95% CI). Climate data indicated anomalously high precipitation in 2014-19 preceding WNV transmission. Data analyzed herein indicate utility in year-round continuous and diversified surveillance methodologies. This information is needed to properly calibrate future models that could assist with predicting transmission events of WNV in Panama City Beach, FL.

Effect of Agroecosystems on Seroprevalence of St. Louis Encephalitis and West Nile Viruses in Birds, La Pampa, Argentina, 2017-2019.

In Argentina, the Pampa ecoregion has been almost completely transformed into agroecosystems. To evaluate the environmental (agricultural area, tree coverage, distance to the nearest water body and urban site) and biological (dove, cowbird, and sparrow abundance) effects on free-ranging bird exposure to St. Louis encephalitis virus (SLEV) and West Nile virus (WNV), we used generalized linear mixed models. For 1,019 birds sampled during 2017-2019, neutralizing antibodies were found against SLEV in samples from 60 (5.8%) birds and against WNV for 21 (2.1%). The best variable for explaining SLEV seroprevalence was agricultural area, which had a positive effect; however, for WNV, no model was conclusive. Our results suggest that agroecosystems in the La Pampa ecoregion increase the exposure of avian hosts to SLEV, thus potentially increasing virus activity.

Impact of climate change on West Nile virus distribution in South America.

BACKGROUND: West Nile virus (WNV) is a vector-borne pathogen of global relevance and is currently the most widely distributed flavivirus causing encephalitis worldwide. Climate conditions have direct and indirect impacts on vector abundance and virus dynamics within the mosquito. The significance of environmental variables as drivers in WNV epidemiology is increasing under the current climate change scenario. In this study we used a machine learning algorithm to model WNV distributions in South America. METHODS: Our model evaluated eight environmental variables for their contribution to the occurrence of WNV since its introduction in South America in 2004. RESULTS: Our results showed that environmental variables can directly alter the occurrence of WNV, with lower precipitation and higher temperatures associated with increased virus incidence. High-risk areas may be modified in the coming years, becoming more evident with high greenhouse gas emission levels. Countries such as Bolivia, Paraguay and several Brazilian areas, mainly in the northeast and midwest regions and the Pantanal biome, will be greatly affected, drastically changing the current WNV distribution. CONCLUSIONS: Understanding the linkages between climatological and ecological change as determinants of disease emergence and redistribution will help optimize preventive strategies. Increased virus surveillance, integrated modelling and the use of geographically based data systems will provide more anticipatory measures by the scientific community.

Domestic Dogs as Sentinels for West Nile Virus but not Aedes-borne Flaviviruses, Mexico.

We tested 294 domestic pet dogs in Mexico for neutralizing antibodies for mosquito-borne flaviviruses. We found high (42.6%) exposure to West Nile virus in Reynosa (northern Mexico) and low (1.2%) exposure in Tuxtla Gutierrez (southern Mexico) but very limited exposure to Aedes-borne flaviviruses. Domestic dogs may be useful sentinels for West Nile virus.

Equids' Core Vaccines Guidelines in North America: Considerations and Prospective.

Vaccination against infectious diseases is a cornerstone of veterinary medicine in the prevention of disease transmission, illness severity, and often death in animals. In North American equine medicine, equine vaccines protecting against tetanus, rabies, Eastern and Western equine encephalomyelitis, and West Nile are core vaccines as these have been classified as having a heightened risk of mortality, infectiousness, and endemic status. Some guidelines differ from the label of vaccines, to improve the protection of patients or to decrease the unnecessary administration to reduce potential side effects. In North America, resources for the equine practitioners are available on the American Association of Equine Practitioners (AAEP) website. Conversely, in small companion animals, peer review materials are regularly published in open access journals to guide the vaccination of dogs and cats. The aims of this review are to present how the vaccine guidelines have been established for small companion animals and horses in North America, to review the equine literature to solidify or contrast the current AAEP guidelines of core vaccines, and to suggest future research directions in the equine vaccine field considering small companion animal strategies and the current available resources in equine literature.

West Nile Virus in Brazil.

Background: West Nile virus (WNV) was first sequenced in Brazil in 2019, when it was isolated from a horse in the Espírito Santo state. Despite multiple studies reporting serological evidence suggestive of past circulation since 2004, WNV remains a low priority for surveillance and public health, such that much is still unknown about its genomic diversity, evolution, and transmission in the country. Methods: A combination of diagnostic assays, nanopore sequencing, phylogenetic inference, and epidemiological modeling are here used to provide a holistic overview of what is known about WNV in Brazil. Results: We report new genetic evidence of WNV circulation in southern (Minas Gerais, São Paulo) and northeastern (Piauí) states isolated from equine red blood cells. A novel, climate-informed theoretical perspective of the potential transmission of WNV across the country highlights the state of Piauí as particularly relevant for WNV epidemiology in Brazil, although it does not reject possible circulation in other states. Conclusion: Our output demonstrates the scarceness of existing data, and that although there is sufficient evidence for the circulation and persistence of the virus, much is still unknown on its local evolution, epidemiology, and activity. We advocate for a shift to active surveillance, to ensure adequate preparedness for future epidemics with spill-over potential to humans.

West Nile Virus in the State of Ceará, Northeast Brazil.

In June 2019, a horse with neurological disorder was diagnosed with West Nile virus (WNV) in Boa Viagem, a municipality in the state of Ceará, northeast Brazil. A multi-institutional task force coordinated by the Brazilian Ministry of Health was deployed to the area for case investigation. A total of 513 biological samples from 78 humans, 157 domestic animals and 278 free-ranging wild birds, as well as 853 adult mosquitoes of 22 species were tested for WNV by highly specific serological and/or molecular tests. No active circulation of WNV was detected in vertebrates or mosquitoes by molecular methods. Previous exposure to WNV was confirmed by seroconversion in domestic birds and by the detection of specific neutralizing antibodies in 44% (11/25) of equids, 20.9% (14/67) of domestic birds, 4.7% (13/278) of free-ranging wild birds, 2.6% (2/78) of humans, and 1.5% (1/65) of small ruminants. Results indicate that not only equines but also humans and different species of domestic animals and wild birds were locally exposed to WNV. The detection of neutralizing antibodies for WNV in free-ranging individuals of abundant passerine species suggests that birds commonly found in the region may have been involved as amplifying hosts in local transmission cycles of WNV.

Ecologic Determinants of West Nile Virus Seroprevalence among Equids, Brazil.

Among 713 equids sampled in northeastern Brazil during 2013-2018, West Nile virus seroprevalence was 4.5% (95% CI 3.1%-6.3%). Mathematical modeling substantiated higher seroprevalence adjacent to an avian migratory route and in areas characterized by forest loss, implying increased risk for zoonotic infections in disturbed areas.

Serosurvey of Nonhuman Primates in Costa Rica at the Human-Wildlife Interface Reveals High Exposure to Flaviviruses.

Arthropod-borne viruses belonging to the flavivirus genus possess an enormous relevance in public health. Neotropical non-human primates (NPs) have been proposed to be susceptible to flavivirus infections due to their arboreal and diurnal habits, their genetic similarity to humans, and their relative closeness to humans. However, the only known flavivirus in the American continent maintained by sylvatic cycles involving NPs is yellow fever virus (YFV), and NPs' role as potential hosts of other flaviviruses is still unknown. Here, we examined flavivirus exposure in 86 serum samples including 83.7% samples from free-range and 16.3% from captive NPs living in flavivirus-endemic regions of Costa Rica. Serum samples were opportunistically collected throughout Costa Rica in 2000-2015. We used a highly specific micro-plaque reduction neutralization test (micro-PRNT) to determine the presence of antibodies against YFV, dengue virus 1-4 (DENV), Zika virus, West Nile virus (WNV), and Saint Louis encephalitis virus (SLEV). We found evidence of seropositive NPs with homotypic reactivity to SLEV 11.6% (10/86), DENV 10.5% (9/86), and WNV 2.3% (2/86). Heterotypic reactivity was determined in 3.5% (3/86) of individuals against DENV, 1.2% (1/86) against SLEV, and 1.2% (1/86) against WNV. We found that 13.9% (12/86) of NPs were positive for an undetermined flavivirus species. No antibodies against DENV-3, DENV-4, YFV, or ZIKV were found. This work provides compelling serological evidence of flavivirus exposure in Costa Rican NPs, in particular to DENV, SLEV, and WNV. The range of years of sampling and the region from where positives were detected coincide with those in which peaks of DENV in human populations were registered, suggesting bidirectional exposure due to human-wildlife contact or bridging vectors. Our work suggests the continuous exposure of wildlife populations to various flaviviruses of public health importance and underscores the necessity of further surveillance of flaviviruses at the human-wildlife interface in Central America.