Recent progress on gene-deleted live-attenuated African swine fever virus vaccines.

African Swine Fever (ASF) is a highly lethal viral disease in swine, with mortality rates approaching 100%. The disease has spread to many swine-producing countries, leading to significant economic losses and adversely impacting global food security. Extensive efforts have been directed toward developing effective ASF vaccines. Among the vaccinology approaches tested to date, live-attenuated virus (LAV) vaccines produced by rational deleting virulence genes from virulent African Swine Fever Virus (ASFV) strains have demonstrated promising safety and efficacy in experimental and field conditions. Many gene-deleted LAV vaccine candidates have been generated in recent years. The virulence genes targeted for deletion from the genome of virulent ASFV strains can be categorized into four groups: Genes implicated in viral genome replication and transcription, genes from the multigene family located at both 5' and 3' termini, genes participating in mediating hemadsorption and putative cellular attachment factors, and novel genes with no known functions. Some promising LAV vaccine candidates are generated by deleting a single viral virulence gene, whereas others are generated by simultaneously deleting multiple genes. This article summarizes the recent progress in developing and characterizing gene-deleted LAV vaccine candidates.

Comparative Analysis of Swine Antibody Responses following Vaccination with Live-Attenuated and Killed African Swine Fever Virus Vaccines.

African swine fever virus (ASFV) is circulating in many swine-producing countries, causing significant economic losses. It is observed that pigs experimentally vaccinated with a live-attenuated virus (LAV) but not a killed virus (KV) vaccine develop solid homologous protective immunity. The objective of this study was to comparatively analyze antibody profiles between pigs vaccinated with an LAV vaccine and those vaccinated with a KV vaccine to identify potential markers of vaccine-induced protection. Thirty ASFV seronegative pigs were divided into three groups: Group 1 received a single dose of an experimental LAV, Group 2 received two doses of an experimental KV vaccine, and Group 3 was kept as a non-vaccinated (NV) control. At 42 days post-vaccination, all pigs were challenged with the parental virulent ASFV strain and monitored for 21 days. All pigs vaccinated with the LAV vaccine survived the challenge. In contrast, eight pigs from the KV group and seven pigs from the NV group died within 14 days post-challenge. Serum samples collected on 41 days post-vaccination were analyzed for their reactivity against a panel of 29 viral structural proteins. The sera of pigs from the LAV group exhibited a strong antibody reactivity against various viral structural proteins, while the sera of pigs in the KV group only displayed weak antibody reactivity against the inner envelope (p32, p54, p12). There was a negative correlation between the intensity of antibody reactivity against five ASFV antigens, namely p12, p14, p15, p32, and pD205R, and the viral DNA titers in the blood of animals after the challenge infection. Thus, antibody reactivities against these five antigens warrant further evaluation as potential indicators of vaccine-induced protection.

Infection of Feral Phenotype Swine with Japanese Encephalitis Virus.

Background: Japanese encephalitis virus (JEV) is a mosquito-borne zoonotic flavivirus and the leading cause of pediatric encephalitis in the Asian Pacific region. The transmission cycle primarily involves Culex spp. mosquitoes and Ardeid birds, with domestic pigs (Sus scrofa domestica) being the source of infectious viruses for the spillover of JEV from the natural endemic transmission cycle into the human population. Although many studies have concluded that domestic pigs play an important role in the transmission cycle of JEV, and infection of humans, the role of feral pigs in the transmission of JEV remains unclear. Since domestic and feral pigs are the same species, and because feral pig populations in the United States are increasing and expanding geographically, the current study aimed to test the hypothesis that if JEV were introduced into the United States, feral pigs might play a role in the transmission cycle. Materials and Methods: Sinclair miniature pigs, that exhibit the feral phenotype, were intradermally inoculated with JEV genotype Ib. These pigs were derived from crossing miniature domestic pig with four strains of feral pigs and were used since obtaining feral swine was not possible. Results: The Sinclair miniature pigs became viremic and displayed pathological outcomes similar to those observed in domestic swine. Conclusion: Based on these findings, we conclude that in the event of JEV being introduced into the United States, feral pig populations could contribute to establishment and maintenance of a transmission cycle of JEV and could lead to the virus becoming endemic in the United States.

Serological Prevalence of Crimean-Congo Hemorrhagic Fever Virus Infection in Small Ruminants and Cattle in The Gambia.

Crimean-Congo hemorrhagic fever virus (CCHFV) is a widely distributed tickborne zoonotic agent that infects a variety of host species. There is a lack of information on the true geographic distribution of the prevalence and risk of CCHFV in West Africa. A countrywide cross-sectional study involving 1413 extensively managed indigenous small ruminants and cattle at livestock sales markets and in village herds, respectively, was carried out in The Gambia. In sheep, an overall anti-CCHFV antibody prevalence of 18.9% (95% CI: 15.5-22.8%), goats 9.0% (95% CI: 6.7-11.7%), and cattle 59.9% (95% CI: 54.9-64.7%) was detected. Significant variation (p < 0.05) in the prevalence of anti-CCHFV antibodies at sites in the five administrative regions (sheep: 4.8-25.9%; goats: 1.8-17.1%) and three agroecological zones (sheep: 8.9-32.9%; goats: 4.1-18.0%) was also observed. Comparatively, higher anti-CCHFV antibody prevalence was detected in cattle (33.3-84.0%) compared to small ruminants (1.8-8.1%). This study represents the first countrywide investigation of the seroprevalence of CCHFV in The Gambia, and the results suggest potential circulation and endemicity of the virus in the country. These data provide critical information vital to the development of informed policies for the surveillance, diagnosis, and control of CCFHV infection in The Gambia and the region.

Preliminary Evaluation of a Recombinant Rift Valley Fever Virus Glycoprotein Subunit Vaccine Providing Full Protection against Heterologous Virulent Challenge in Cattle.

Rift Valley fever virus (RVFV) is a mosquito-borne zoonotic pathogen that causes periodic outbreaks of abortion in ruminant species and hemorrhagic disease in humans in sub-Saharan Africa. These outbreaks have a significant impact on veterinary and public health. Its introduction to the Arabian Peninsula in 2003 raised concerns of further spread of this transboundary pathogen to non-endemic areas. These concerns are supported by the presence of competent vectors in many non-endemic countries. There is no licensed RVF vaccine available for humans and only a conditionally licensed veterinary vaccine available in the United States. Currently employed modified live attenuated virus vaccines in endemic countries lack the ability for differentiating infected from vaccinated animals (DIVA). Previously, the efficacy of a recombinant subunit vaccine based on the RVFV Gn and Gc glycoproteins, derived from the 1977 human RVFV isolate ZH548, was demonstrated in sheep. In the current study, cattle were vaccinated subcutaneously with the Gn only, or Gn and Gc combined, with either one or two doses of the vaccine and then subjected to heterologous virus challenge with the virulent Kenya-128B-15 RVFV strain, isolated from Aedes mosquitoes in 2006. The elicited immune responses by some vaccine formulations (one or two vaccinations) conferred complete protection from RVF within 35 days after the first vaccination. Vaccines given 35 days prior to RVFV challenge prevented viremia, fever and RVFV-associated histopathological lesions. This study indicates that a recombinant RVFV glycoprotein-based subunit vaccine platform is able to prevent and control RVFV infections in target animals.

Management Strategies for Reducing the Risk of Equines Contracting Vesicular Stomatitis Virus (VSV) in the Western United States.

Vesicular stomatitis viruses (VSVs) cause a condition known as vesicular stomatitis (VS), which results in painful lesions in equines, cattle, swine, and camelids, and when transmitted to humans, can cause flu-like symptoms. When animal premises are affected by VS, they are subject to a quarantine. The equine industry more broadly may incur economic losses due to interruptions of animal trade and transportation to shows, competitions, and other events. Equine owners, barn managers, and veterinarians can take proactive measures to reduce the risk of equines contracting VS. To identify appropriate risk management strategies, it helps to understand which biting insects are capable of transmitting the virus to animals, and to identify these insect vectors' preferred habitats and behaviors. We make this area of science more accessible to equine owners, barn managers, and veterinarians, by (1) translating the most relevant scientific information about biting insect vectors of VSV and (2) identifying practical management strategies that might reduce the risk of equines contracting VSV from infectious biting insects or from other equines already infected with VSV. We address transmission risk at four different spatial scales-the animal, the barn/shelter, the barnyard/premises, and the surrounding environment/neighborhood-noting that a multiscale and spatially collaborative strategy may be needed to reduce the risk of VS.

Perspectives Regarding the Risk of Introduction of the Japanese Encephalitis Virus (JEV) in the United States.

Japanese encephalitis (JE) is a zoonotic, emerging disease transmitted by mosquito vectors infected with the Japanese encephalitis virus (JEV). Its potential for emergence into susceptible regions is high, including in the United States (US), and is a reason of economic concern among the agricultural community, and to public health due to high morbidity and mortality rates in humans. While exploring the complexities of interactions involved with viral transmission, we proposed a new outlook on the role of vectors, hosts and the environment under changing conditions. For instance, the role of feral pigs may have been underappreciated in our previous work, given research keeps pointing to the importance of susceptible populations of wild swine in naïve regions as key elements for the introduction of emergent vector-borne diseases. High risk of JEV introduction has been associated with the transportation of infected mosquitoes via aircraft. Nonetheless, no JEV outbreaks have been reported in the US to date and results from a qualitative risk assessment considered the risk of establishment to be negligible under the current conditions (environmental, vector, pathogen, and host). In this work, we discuss virus-vector-host interactions and ecological factors important for virus transmission and spread, review research on the risk of JEV introduction to the US considering the implications of risk dismissal as it relates to past experiences with similar arboviruses, and reflect on future directions, challenges, and implications of a JEV incursion.

Evaluation of A Baculovirus-Expressed VP2 Subunit Vaccine for the Protection of White-Tailed Deer (Odocoileus virginianus) from Epizootic Hemorrhagic Disease.

Epizootic hemorrhagic disease virus (EHDV) is an arthropod-transmitted RNA virus and the causative agent of epizootic hemorrhagic disease (EHD) in wild and domestic ruminants. In North America, white-tailed deer (WTD) experience the highest EHD-related morbidity and mortality, although clinical disease is reported in cattle during severe epizootics. No commercially licensed EHDV vaccine is available in North America. The objective of this study was to develop and evaluate a subunit vaccine candidate to control EHD in WTD. Recombinant VP2 (rVP2) outer capsid proteins of EHDV serotypes 2 (EHDV-2) and 6 (EHDV-6) were produced in a baculovirus-expression system. Mice and cattle vaccinated with EHDV-2 or EHDV-6 rVP2 produced homologous virus-neutralizing antibodies. In an immunogenicity/efficacy study, captive-bred WTD received 2 doses of EHDV-2 rVP2 or sham vaccine, then were challenged with wild-type EHDV-2 at 30 d post vaccination. None of the rVP2-vaccinated deer developed clinical disease, no viral RNA was detected in their blood or tissues (liver, lung, spleen, kidney), and no EHDV-induced lesions were observed. Sham-vaccinated deer developed clinical disease with viremia and typical EHD vascular lesions. Here, we demonstrate a rVP2 subunit vaccine that can provide protective immunity from EHDV infection and which may serve as an effective tool in preventing clinical EHD and reducing virus transmission.

Immunogenicity and efficacy of Schmallenberg virus envelope glycoprotein subunit vaccines.

The Schmallenberg virus (SBV) is an orthobunyavirus that causes abortions, stillbirths, and congenital defects in pregnant sheep and cattle. Inactivated or live attenuated vaccines have been developed in endemic countries, but there is still interest in the development of SBV vaccines that would allow Differentiating Infected from Vaccinated Animals (DIVA). Therefore, an attempt was made to develop novel DIVA-compatible SBV vaccines using SBV glycoproteins expressed in baculovirus. All vaccines and phosphate buffered saline (PBS) controls were prepared with adjuvant and administered subcutaneously to cattle at 6 month of age. The first trial included 2 groups of animals vaccinated with either carboxyl-terminus glycoprotein (Gc) or PBS and boosted after 2 weeks. In the second trial, 3 groups of cattle were administered either Gc, Gc and amino-terminus glycoprotein (Gn), or PBS with a booster vaccination after 3 weeks. The animals were challenged with SBV 9 days after the booster vaccination in the first study, and 3 weeks after the booster vaccination in the second study. Using a SBV Gc-specific enzyme-linked immunosorbent assay, antibodies were first detected in serum samples 14 days after the first vaccination in both trials, and peaked on days 7 and 9 after the booster in the first and second trials, respectively. Low titers of neutralizing antibodies were detected in serum from only 3/6 and 2/4 animals in the first and second trial, respectively, at 14 days after the first vaccination. The titers increased 2 to 3-fold after the booster vaccination. SBV-specific RNA was detected in the serum and selective tissues in all animals after SBV challenge independent of vaccination status. The SBV candidate vaccines neither prevented viremia nor conferred protection against SBV infection.

Evaluation of an Indirect Enzyme-Linked Immunosorbent Assay Based on Recombinant Baculovirus-Expressed Rift Valley Fever Virus Nucleoprotein as the Diagnostic Antigen.

The increasing risk of Rift Valley fever virus (RVFV) infection as a global veterinary and public health threat demands the development of safe and accurate diagnostic tests. The aim of this study was to assess the suitability of a baculovirus expression system to produce recombinant RVFV nucleoprotein (N) for use as serodiagnostic antigen in an indirect enzyme-linked immunosorbent assay (ELISA). The ability of the recombinant N antigen to detect RVFV antibody responses was evaluated in ELISA format using antisera from sheep and cattle experimentally infected with two genetically distinct wild-type RVFV strains and sera from indigenous sheep and goat populations exposed to natural RVFV field infection in The Gambia. The recombinant N exhibited specific reactivity with the N-specific monoclonal antibody and various hyperimmune serum samples from ruminants. The indirect ELISA detected N-specific antibody responses in animals with 100% sensitivity compared to the plaque reduction neutralization test (6 to 21 days postinfection) and with 97% and 100% specificity in sheep and cattle, respectively. There was a high level of correlation between the indirect N ELISA and the virus neutralization test for sheep sera (R2 = 0.75; 95% confidence interval [CI] = 0.73 to 0.92) and cattle sera (R2 = 0.80; 95% CI = 0.67 to 0.97); in addition, the N-specific ELISA detected RVFV seroprevalence levels of 26.1% and 54.3% in indigenous sheep and goats, respectively, in The Gambia. The high specificity and correlation with the virus neutralization test support the idea of the feasibility of using the recombinant baculovirus-expressed RVFV N-based indirect ELISA to assess RVFV seroprevalence in livestock in areas of endemicity and nonendemicity.

EHDV-2 Infection Prevalence Varies in Culicoides sonorensis after Feeding on Infected White-Tailed Deer over the Course of Viremia.

Epizootic hemorrhagic disease viruses (EHDVs) are arboviral pathogens of white-tailed deer and other wild and domestic ruminants in North America. Transmitted by various species of Culicoides, EHDVs circulate wherever competent vectors and susceptible ruminant host populations co-exist. The impact of variation in the level and duration of EHDV viremia in white-tailed deer (Odocoileus virginianus) on Culicoides infection prevalence is not well characterized. Here we examined how infection prevalence in a confirmed North American vector of EHDV-2 (Culicoides sonorensis) varies in response to fluctuations in deer viremia. To accomplish this, five white-tailed deer were experimentally infected with EHDV-2 and colonized C. sonorensis were allowed to feed on deer at 3, 5, 7, 10, 12, 14, 18, and 24 days post infection (dpi). Viremia profiles in deer were determined by virus isolation and titration at the same time points. Blood-fed Culicoides were assayed for virus after a 10-day incubation (27 °C) period. We found that increases in deer EHDV blood titers significantly increased both the likelihood that midges would successfully acquire EHDV and the proportion of midges that reached the titer threshold for transmission competence. Unexpectedly, we identified four infected midge samples (three individuals and one pool) after feeding on one deer 18 and 24 dpi, when viremia was no longer detectable by virus isolation. The ability of ruminants with low-titer viremia to serve as a source of EHDV for blood-feeding Culicoides should be explored further to better understand its potential epidemiological significance.

A spatio-temporal individual-based network framework for West Nile virus in the USA: Spreading pattern of West Nile virus.

West Nile virus (WNV)-a mosquito-borne arbovirus-entered the USA through New York City in 1999 and spread to the contiguous USA within three years while transitioning from epidemic outbreaks to endemic transmission. The virus is transmitted by vector competent mosquitoes and maintained in the avian populations. WNV spatial distribution is mainly determined by the movement of residential and migratory avian populations. We developed an individual-level heterogeneous network framework across the USA with the goal of understanding the long-range spatial distribution of WNV. To this end, we proposed three distance dispersal kernels model: 1) exponential-short-range dispersal, 2) power-law-long-range dispersal in all directions, and 3) power-law biased by flyway direction -long-range dispersal only along established migratory routes. To select the appropriate dispersal kernel we used the human case data and adopted a model selection framework based on approximate Bayesian computation with sequential Monte Carlo sampling (ABC-SMC). From estimated parameters, we find that the power-law biased by flyway direction kernel is the best kernel to fit WNV human case data, supporting the hypothesis of long-range WNV transmission is mainly along the migratory bird flyways. Through extensive simulation from 2014 to 2016, we proposed and tested hypothetical mitigation strategies and found that mosquito population reduction in the infected states and neighboring states is potentially cost-effective.

Introduction of the Japanese encephalitis virus (JEV) in the United States - A qualitative risk assessment.

The purpose of this risk assessment (RA) was to qualitatively estimate the risk of emergence of the Japanese encephalitis virus (JEV) in the United States (US). We followed the framework for RA of emerging vector-borne livestock diseases (de Vos et al. 2011), which consists of a structured questionnaire, whose answers to questions can be delivered in risk categories, descriptive statements, or yes or no type of answers, being supported by the literature. The most likely pathways of introduction of JEV identified were: (a) entry through infected vectors (by aircraft, cargo ships, tires, or wind); (b) import of infected viremic animals; (c) entry of viremic migratory birds; (d) import of infected biological materials; (e) import of infected animal products; (f) entry of infected humans; and (g) import/production of contaminated biological material (e.g., vaccines). From these pathways, the probability of introduction of JEV through infected adult mosquitoes via aircraft was considered very high and via ships/containers was deemed low to moderate. The probability of introduction via other pathways or modes of entry (vector eggs or larvae, hosts, and vaccines) was considered negligible. The probability of transmission of JEV was variable, ranging from low to high (in the presence of both competent vectors and hosts), depending on the area of introduction within the US. Lastly, the probability of establishment of JEV in the continental US was considered negligible. For that reason, we stopped the risk assessment at this point of the framework. This RA provides important information regarding the elements that contribute to the risk associated with the introduction of JEV in the US. This RA also indicates that infected mosquitoes transported in aircraft (and cargo ships) are the most likely pathway of JEV entry and therefore, mitigation strategies should be directed towards this pathway.

A quantitative risk assessment (QRA) of the risk of introduction of the Japanese encephalitis virus (JEV) in the United States via infected mosquitoes transported in aircraft and cargo ships.

Following a qualitative risk assessment, in which we identified and assessed all viable pathways for the introduction of the Japanese encephalitis virus (JEV) into the United States (US), we identified entry through infected vectors via aircraft and cargo ships as the most likely pathway, and thus considered it further in a quantitative risk assessment (QRA) model. The objective of this study was to evaluate the risk of introduction of JEV in the US via infected mosquitoes transported in aircraft and cargo ships arriving from Asia, using a QRA model. We created a stochastic model to quantify the probability of introduction of at least one infected mosquito in the continental US via aircraft and cargo ships, per at-risk period (March to October) or year, respectively. We modeled the following parameters: number of flights (per at-risk period, i.e., March to October) and cargo ships (per year) and per region, number of mosquitoes per flight and ship, number of mosquitoes that were not found and sensitivity of the mosquito collection method in aircraft, mosquito infection rates, and number of mosquitoes coming in aircraft per at-risk period (March to October) and cargo ships per year. Flight and cargo ship data pertained to years 2010-2016. For model building purposes, we only considered port-to-port vessels arriving from Asia to the US, we assumed that mosquitoes survive the trans-Pacific Ocean ship crossing and that the number of mosquitoes in cargo and passenger flights is similar. Our model predicted a very high risk (0.95 median probability; 95% CI = 0.80-0.99) of at least one infected mosquito being introduced in the US during the at-risk period, i.e., March to October, via aircraft transportation from JEV-affected countries in Asia. We also estimated that a median of three infected mosquitoes can enter the US during the at-risk period, i.e., March to October (95% CI = 1-7). The highest probability of introduction via aircraft was attributed to the Mediterranean California ecoregion (0.74; 95% CI = 0.50-0.90). We predicted, however, a negligible risk (0; 95% CI = 0.00-0.01) of at least one infected mosquito being introduced via cargo ships. Although the risk of introduction of JEV-infected mosquitoes by cargo ships was negligible, the risk via aircraft was estimated to be high. Our findings indicate the need to prioritize JEV prevention and control methods for aircraft-based pathways, such as aircraft disinfection. The quantitative estimates provided in this study are of interest to public health entities and other stakeholders, as they may support future interventions for preventing JEV introduction, as well as other vector-borne diseases, in the US and other countries.

Susceptibility of White-Tailed Deer to Rift Valley Fever Virus.

Rift Valley fever virus, a zoonotic arbovirus, poses major health threats to livestock and humans if introduced into the United States. White-tailed deer, which are abundant throughout the country, might be sentinel animals for arboviruses. We determined the susceptibility of these deer to this virus and provide evidence for a potentially major epidemiologic role.

Virological and Serological Responses of Sheep and Cattle to Experimental Schmallenberg Virus Infection.

Schmallenberg virus (SBV) is an orthobunyavirus in the Simbu serogroup that emerged in Germany in late 2011 and was mostly associated with a mild transient disease of sheep and cattle. SBV is transmitted by biting midges (Culicoides species) and causes abortions, stillbirths, and congenital defects in naïve pregnant ruminants. Two separate studies were conducted with a primary objective of better understanding the virological and serological responses of sheep and cattle to different SBV isolates after experimental infection. The second objective was to produce immunoreagents and challenge materials for use in future vaccine and diagnostics research. These studies were carried out using the following infectious inocula: (i) infectious serum (IS) (ii) cell culture-grown virus, and (iii) infectious lamb brain homogenate. The responses were assessed in both species throughout the course of the experiment. SBV RNA in serum (RNAemia) was detected as early as 2 (in sheep) and 3 (in cattle) days postinfection (dpi) and peaked on 3 and 4 dpi in cattle and sheep, respectively. Cattle had higher levels of RNAemia compared with sheep. Experimental infection with IS resulted in the highest level of RNAemia in both species followed by cell culture-grown virus. A delayed, low level RNAemia was detected in cattle inoculated with infectious sheep brain. Isolation of SBV was only possible from 4 dpi sera from all cattle inoculated with IS and one sheep inoculated with cell culture-derived virus. SBV neutralizing antibodies were first detected on 14 dpi in both species. No specific gross and microscopic lesions were observed in either study. In conclusion, these studies highlight not only the difference in viremia and anti-SBV antibody level against the different SBV isolates, but also the extent of the response in the two host species.

Physiological and immunological responses to Culicoides sonorensis blood-feeding: a murine model.

BACKGROUND: Hematophagous Culicoides spp. biting midges are of great agricultural importance as livestock, equine, and wildlife pests and as vectors of the orbiviruses bluetongue, epizootic hemorrhagic disease and African horse sickness. To obtain a blood meal, midges deposit saliva containing allergens, proteases, and anti-hemostatic factors, into the dermis to facilitate feeding. Infected midges deposit virus along with the myriad of salivary proteins during feeding. The extreme efficiency with which midges are able to transmit orbiviruses is not clearly understood, as much is still unknown about the physiological trauma of the bite and immune responses to saliva deposited during feeding. Of particular interest are the first few hours and days after the bite; a critical time period for any midge-transmitted virus to quickly establish a localized infection and disseminate, while avoiding the hosts' immune responses. RESULTS: A mouse-midge feeding model using colonized Culicoides sonorensis midges was used to characterize innate mammalian immune responses to blood-feeding. Histological analysis of skin, and cellular and cytokine profiles of draining lymph nodes show Culicoides midge feeding elicited a potent pro-inflammatory Th-mediated cellular response with significant mast cell activation, subcutaneous hematomas, hypodermal edema and dermal capillary vasodilation, and rapid infiltration of leukocytes to the bite sites. Mast cell degranulation, triggered by bite trauma and specifically by midge saliva, was key to physiological and immunological responses and the ability of midges to feed to repletion. CONCLUSIONS: Midge feeding causes physiological and immunological responses that would be highly favorable for rapid infection and systemic dissemination orbiviruses if delivered during blood-feeding. Recruitment of leukocytic cells to bitten skin brings susceptible cell populations in proximity of deposited virus within hours of feeding. Infected cells would drain to lymph nodes, which become hyperplastic in response to saliva, and result in robust viral replication in expanding cell populations and dissemination via the lymph system. Additionally, saliva-induced vasodilation and direct breaches in dermal capillaries by biting mouthparts exposes susceptible vascular endothelial cells, thereby providing immediate sites of virus replication and a dissemination route via the circulatory system. This research provides insights into the efficiency of Culicoides midges as orbivirus vectors.

North American domestic pigs are susceptible to experimental infection with Japanese encephalitis virus.

Japanese encephalitis virus (JEV) is a mosquito-borne flavivirus that is capable of causing encephalitic diseases in children. While humans can succumb to severe disease, the transmission cycle is maintained by viremic birds and pigs in endemic regions. Although JEV is regarded as a significant threat to the United States (U.S.), the susceptibility of domestic swine to JEV infection has not been evaluated. In this study, domestic pigs from North America were intravenously challenged with JEV to characterize the pathological outcomes. Systemic infection followed by the development of neutralizing antibodies were observed in all challenged animals. While most clinical signs were limited to nonspecific symptoms, virus dissemination and neuroinvasion was observed at the acute phase of infection. Detection of infectious viruses in nasal secretions suggest infected animals are likely to promote the vector-free transmission of JEV. Viral RNA present in tonsils at 28 days post infection demonstrates the likelihood of persistent infection. In summary, our findings indicate that domestic pigs can potentially become amplification hosts in the event of an introduction of JEV into the U.S. Vector-free transmission to immunologically naïve vertebrate hosts is also likely through nasal shedding of infectious viruses.

High-Polyphenol Sorghum Bran Extract Inhibits Cancer Cell Growth Through ROS Induction, Cell Cycle Arrest, and Apoptosis.

As diet is one of the major controllable factors in cancer development, potentially chemopreventive foods are of significant interest to public health. One such food is sorghum (Sorghum bicolor), a cereal grain that contains varying concentrations of polyphenols. In a panel of 15 sorghum germplasm, we identified strains with higher polyphenol content than previously reported for this grain. Bran extracts from the germplasm with the highest and lowest polyphenol content were then tested against HepG2 and Caco2 cancer cells to assess effects on cancer cell viability, reactive oxygen species, apoptosis, DNA damage, cell cycle arrest, and protein expression patterns. High-polyphenol extracts, but not low-polyphenol extracts, reduced cell viability by inducing apoptosis and cell cycle arrest following production of reactive oxygen species and oxidative DNA damage. The results indicate that high-polyphenol sorghum bran extracts have potential anticancer properties and warrant further research, not only to test against specific cancers but also to elucidate underlying mechanisms of action.

Shedding of Japanese Encephalitis Virus in Oral Fluid of Infected Swine.

Japanese encephalitis virus (JEV) is a zoonotic mosquito-borne flavivirus endemic in the Asia-Pacific region. Maintenance of JEV in nature involves enzootic transmission by competent Culex mosquitoes among susceptible avian and swine species. Historically, JEV has been regarded as one of the most important arthropod-borne viruses in Southeast Asia. Oronasal shedding of JEV from infected amplification hosts was not recognized until the recent discovery of vector-free transmission of JEV among domestic pigs. In this study, oral shedding of JEV was characterized in domestic pigs and miniature swine representing the feral phenotype. A rope-based sampling method followed by the detection of viral RNA using RT-qPCR allowed the collection and detection of JEV in oral fluid samples collected from intradermally challenged animals. The results suggest that the shedding of JEV in oral fluid can be readily detected by molecular diagnostic assays at the acute phase of infection. It also demonstrates the feasibility of this technique for the diagnosis and surveillance of JEV in swine species.