Many novel genetic approaches are under development to combat insect pests. One genetic strategy aims to suppress or locally eliminate a species through large, repeated releases of genetically engineered strains that render female offspring unviable under field conditions. Strains with this female‐killing characteristic have been developed either with all of the molecular components in a single construct or with the components in two constructs inserted at independently assorting loci. Strains with two constructs are typically considered to be only of value as research tools and for producing solely male offspring in rearing factories which are subsequently sterilized by radiation before release. A concern with the two‐construct strains is that once released, the two constructs would become separated and therefore non‐functional. The only female‐killing strains that have been released in the field without sterilization are single‐construct strains. Here, we use a population genetics model with density dependence to evaluate the relative effectiveness of female‐killing approaches based on single‐ and two‐construct arrangements. We find that, in general, the single‐construct arrangement results in slightly faster population suppression, but the two‐construct arrangement can eventually cause stronger suppression and cause local elimination with a smaller release size. Based on our results, there is no a priori reason that males carrying two independently segregating constructs need to be sterilized prior to release. In some cases, a fertile release would be more efficient for population suppression. Often this is achieved through a Tet‐off system where tetracycline in the diet represses the activator for a lethal gene  the tetracycline‐repressible transactivator (tTA) with a promoter; and (2) a lethal gene with an enhancer/promoter consisting of multiple tTA binding sites (tetO) and a core promoter. In the initial two‐component systems, tTA was expressed in females by using a female‐specific promoter  and late (adult) expected viabilities for individuals of sex S and genotype i. Tables We assume complete effectiveness of the constructs, so when there is no gene repression via tetracycline, all individuals bearing the functional BK system and all females with the functional FK system die, with a genotype viability of zero . For the 2‐locus system, we allow for unequal fitness costs between each of the insertions. We let two copies of the A allele contribute a fitness cost of sxcA, where cA is the proportion of the total 2‐locus fitness cost accounted for by the A allele, and one copy of the A allele contributes a fitness cost of h·sxcA. A single B allele contributes a fitness cost of h·sx(1‐cA), while being homozygous for B induces a cost of sx(1‐cA). Note that A can be arbitrarily defined as whichever component is more costly because the two alleles only matter in combination. Resulting fitness expressions for all genotypes are listed in Tables We generally assume that the fitness costs are equal for the homozygotes in the 1‐locus and 2‐locus systems to facilitate a direct comparison between the two systems. The 2‐locus system has the same components as the 1‐locus system, which makes equal fitness costs a reasonable base assumption for the purposes of this work  and JiF(t) be the number of juvenile (larvae and pupae) males and females, respectively, of genotype i at time t, and AiM(t) and AiF(t) be the number of viable adult male and adult female mosquitoes in the population, respectively, of genotype i at time t. This gives a maximum of 12 classes of individuals to track  for the 1‐locus system and 36 classes for the 2‐locus system, though lethality from the genetic construct prevents survival of certain classes. For instance, E‐BK only has five nonzero classes . We also assume that in the absence of early sex‐dependent lethality due to transgenes, males and females have equal birth rates and equal hatching fitness costs, allowing a further reduction in the number of unique classes for late‐acting approaches because JiM(t)=JiF(t) for all i and t. This results in seven classes for L‐FK1  and 23 classes for L‐FK2 . These dimensionality reductions can be useful when finding analytical solutions, but for simplicity, we computationally simulated all 12 (1‐locus) or 36 (2‐locus) equations.We model genotype counts over time using a system of ordinary differential equations adapted from Robert et al. . We let i at time t at rateλ is the per‐capita birth rate and Pi|m,n is the probability that a juvenile produced from a mating between a female and male of genotypes m and n, respectively, will be of genotype i. The fraction gives the probability that a randomly chosen male adult is of genotype n, weighted by mating competitiveness wnM. The offspring genotype probabilities are calculated assuming Mendelian inheritance, and for the 2‐locus case, independent segregation of genes at each locus.Accounting for fitness costs, adult females produce juveniles of genotype v. We assume juveniles, adult males, and adult females have per‐capita density‐independent mortality rates of μJ, μM, and μF, respectively. Juveniles also undergo density‐dependent mortality at a per‐capita rate αJ(t)β‐1, where J(t) is the total number of juveniles at time t, and α and β are parameters related to the strength of density‐dependent mortality. The strength of density dependence is adjusted by varying β, with higher β resulting in a faster return to equilibrium population size after a small perturbation. A value of β = 2 gives the logistic model for population dynamics. Given the many complexities and limited understanding of density‐dependent effects in field Ae. aegypti systems /7, where r is the weekly release ratio (engineered:wild‐type) based on the equilibrium number of males prior to the release. By maintaining a constant number of released males, the effective release ratio increases as the population size decreases. The release genotype is KK for 1‐locus (i = 3) and AABB for 2‐locus (i = 9), and because we assume that no females are released, uiF=0 for all i.We assume a continuous release of homozygote engineered males at (daily) rate JiS(t), J(t), AiS(t), and Bi(t) omitted for simplicity of notation) isi = 1 N and S = F or M. All model parameters are listed in Table Ae. aegypti used by Robert et al.  causes the juvenile mortality rates to decrease more quickly as population size decreases from equilibrium. In a system with strong density dependence, the weekly release ratio (r) must be larger to achieve the same amount of population suppression as in a system with weak density dependence. Large r can result in target population extinction, which we define as a time point when there are no females left in the population, even though males are still being released. In contrast, small r results in a new, lower equilibrium population density, where the proportion of individuals that die due to bearing the transgene is not high enough to outweigh the increased survival of juveniles due to decreased density‐dependent mortality in the smaller population.Each of the FK  and BK (bisex killing) genetic strategies has the goal of causing the population to decline by reducing the number of reproductive adult females. The strength of density‐dependent mortality moderates the reductions in population size because stronger density dependence , releases result in weaker suppression of the population, with none of the approaches causing extinction of the population if β = 4.Figure cr, above which the release is large enough to cause the population to go extinct. With such a large number of released males, population extinction is the only stable equilibrium, meaning the release will cause extinction regardless of initial population size. For ongoing release at release ratios below cr, there is a nonzero stable equilibrium for the number of viable adult females, meaning that release will not push a wild‐type population to extinction. A population size of zero is also stable, but the system will approach the nonzero equilibrium unless starting from very low population sizes; that is, it is a bistable (or multistable) system with a low unstable equilibrium  immigration of wild‐type from re‐establishing a population. Mathematical details on the shift in qualitative behavior  at cr, including analysis using Mathematica , and small release ratios result in a suppressed but nonzero equilibrium population size. For most sets of parameters, there is a critical release ratio, β = 3. As a result of the bifurcation, each of the lines is discontinuous, jumping from a nonzero equilibrium to a zero equilibrium at cr. The smallest release size required to cause extinction for each approach is rcLBK = 0.86 < rcLFK2 = 2.52 < rcLFK1 = 2.86 < rcEFK2 = 4.74 < rcEFK1 = 5.40 < rcEBK = 5.97. This order of effectiveness matches that of Figure r is below the critical release ratios and 1‐locus FK requiring a release ratio <15% larger than 2‐locus FK to cause extinction.Figure r = 12, for example, the times are 86 (L‐BK), 114 (L‐FK1), 119 (L‐FK2), 133 (E‐BK), 149 (E‐FK1), and 153 (E‐FK2) days. Note, however, that at lower release ratios, 2‐locus FK systems have a lower time to extinction than 1‐locus FK systems. The results are similar for β = 2 , there is a region with low fitness costs where there is fairly little variation between the time it takes to suppress the population below the threshold. As one or both of the fitness costs increase, there is a margin of longer times separating the successful suppression region from the unsuccessful region. As evident from Figure r = 7 is only slightly greater than cr for those parameter values, and a system with even higher fitness costs has cr > 7, so suppression below 0.05% of equilibrium is not achieved.Results over a wide range of fitness parameters highlight the minimal differences between FK1 and FK2. We use the time it takes to suppress the number of viable females to under 0.05% of the equilibrium when there is a high release ratio of s Figure . For eaccr in most of the region with successful release. Even with different fitness costs, FK1 and FK2 do not differ drastically in time until extinction; FK2 would be substantially less effective only if the costs become large enough for the release to become too small to cause extinction . Unpacking the differences between all approaches, for the E‐BK approach, neither h nor Hs affect the length of time for reduction; only released males experience costs so the degree of dominance does not affect results, and all offspring that inherit a transgene are inviable so hatching costs do not affect results. For E‐BK, increasing the cost to male mating competitiveness drastically decreases release efficacy because doing so effectively reduces the release ratio , and with no surviving transgenic offspring, the release ratio is the main contributing factor to E‐BK efficacy. When Hs = 1 and h = 1, none of the offspring survive for the E‐FK1 and E‐FK2 approaches, making the times shown identical to E‐BK. Decreasing these parameters makes a difference for FK approaches because the males are subject to fitness costs. Dominant fitness costs (bottom row) actually enable successful population reduction for higher male mating costs. With large, dominant mating costs, FK approaches have few mating adult males, becoming effectively similar to the highly effective L‐BK, which has no adult males. The full explanation for increased effectiveness with higher degrees of dominance relates to the propagation of wild‐type alleles, similar to the previous description: homozygote males that are being released are affected regardless of h, and when h > 0, the fitness cost also reduces the spread of wild‐type alleles in heterozygotes. This effect is also evident in other transgenic systems, such as two‐locus underdominance, where dominant transgenic allele fitness costs prevent the wild‐type from being maintained in the population at small frequencies . Additionally, our model does not account for an Allee effect, where there is a critical population size below which the population is unable to maintain itself. With an Allee effect, suppression does not need to be complete to lead to extinction of the population; this may benefit 1‐locus and 2‐locus approaches differently. Finally, given its generality, our model does not account for any potential mechanisms for resistance development. Depending on the mechanism of lethality, there may be advantages for having both components for lethality inserted together. While these areas require further investigation, our results indicate that overall, there is little difference in the pest population suppression efficacies of 1‐locus FK and 2‐locus FK.None declared.Supplementary MaterialClick here for additional data file.


In Brazil, West Nile virus (WNV) was first detected, in 2018, in horses with neurological disease.  We report the first case of WNV infection in a horse from Ceará state and the complete genome sequence of an isolate from Espírito Santo state. Both infections occurred in 2019.  WNV was isolated from the tissues of a horse with neurological signs in Espírito Santo and sequenced by MiSeq.  Phylogenetic analysis revealed that the isolate belongs to lineage 1a, clustering with the NY99 strain, a strain that has not circulated in the USA since 2005.  Our findings reinforce the hypothesis that WNV has been silently circulating in Brazil for many years. Flaviviridae and Flavivirus genera. It belongs to the Japanese encephalitis virus serocomplex that contains other viruses that cause neurological diseases in humans and animals, such as the Usutu, Saint Louis encephalitis, Murray Valley encephalitis, and Cacipacore viruses,Aedes, Ochlerotatus, Anopheles, and Psorophora) in North America, Culex mosquitoes are the main vectors involved in WNV transmission. These mosquitoes are involved in enzootic and epizootic transmissions, where horses and humans are accidental hosts and do not participate in the virus transmission network-,-The West Nile virus (WNV) is a zoonotic RNA virus belonging to the In June 2019, two fatal cases of horses with neurological disease were reported to the Brazilian government, one from Espírito Santo (804_02_ES) and another from Ceará state (827_01_CE) . Clinica®) in DMEM (HycloneTM) supplemented with 2% fetal bovine serum (FBS) and 2% antibiotic-antimycotic (CultilabTM). The homogenate was clarified at 16,000 × g for 10 min at 4 °C and the supernatant was inoculated into VERO (CCL 81) and C6/36 (CRL 1660) cells. These cells were incubated at 37oC and 28oC, respectively, for 1h with gentle rocking every 10 min. The inoculum was then removed, the maintenance medium (2% FBS) was added to the monolayers, and the inoculated cells were incubated again, at 37 °C (VERO) and 28 °C (C6/36). After 5-6 days, cytopathic effects were clearly observed in VERO cells, and WNV was successfully isolated from 804_02_ES samples in both cell types. A second inoculation of both cells was performed to increase the virus titer.The samples were thawed on ice and 0.03-0.05 grams of either spinal cord (804_02_ES) or encephalon (827_01_CE) were manually macerated with a microcentrifuge pestle , Trimmomatic version 0.3.9 (http://www.usadellab.org/cms/?page=trimmomatic) and AfterQC version 0.9.7 (https://github.com/OpenGene/AfterQC). Virus assembly was mapped against the NY99 strain (GenBank accession: MH643887) to obtain a consensus sequence. The final consensus sequence of 804_02_ES was 10,893 nucleotides in length and an identity of 99.71% and 93.44%, for the nucleotide and protein, respectively. We conducted a https://www.viprbrc.org). A total of 116 WNV sequences worldwide (1953-2018) were selected. The dataset was edited and curated for use in the phylogenetic tree to maintain the entire ORF. The Maximum-Likelihood (ML) tree that was created using the IQ-TREE 2.0 software (http://www.iqtree.org) with automatic selection, was GTR+F+I+G4 the best nucleotide substitution model, with a support analysis of 10,000 replicates. The tree graph was edited using iTOL v.5 (https://itol.embl.de).The 804_02_ES genome was aligned using MAFFT software (version 7.0) with a dataset obtained from NCBI and ViPR (The 804_02_ES strain genomic sequence (GenBank accession: MT905060) was phylogenetically determined to belong to WNV lineage 1a  in accorAccording to the Brazilian Ministry of Health, WNV has been officially circulating in Brazil's northeast region since 2014Culex quinquefasciatus) capable of efficient transmission, similar to other Culex species prevalent in the USA? (6) Since WNV has been circulating in Brazil for many years, why has an outbreak of human infections not yet occurred? (7) Is this due to cross-protection induced by other endemic flavivirus infections ? Despite the evidence that WNV has been circulating in Brazil for some time, many questions still remain: (1) How was WNV introduced into Brazil and how has the NY99 signature been preserved until now? (2) Why was it not detected until-2014/2018? (3) Why has no other strain (WN02/SW03) been introduced (or detected) in the country? (4) Which mosquito and bird species are involved in the WNV transmission cycle in the Brazilian territory? (5) Is our most prevalent vector , from the Ministry of Agriculture, Livestock, and Supply (MAPA) to perform the diagnosis. Both animals died naturally. This study does not require approval by the Ethics Committees according to Brazilian normative resolution no. 30 of February 2, 2016, of the National Animal Experimentation Control Council (CONCEA).IFA was performed using hyperimmune serum obtained by venipuncture from infected mice. The Ethics Committee on the Use of Animals (CEUA) from Ribeirão Preto Medical School, University of São Paulo, approved the study under process number 214/2019.


Bandavirus consists of seven tick-borne bunyaviruses, among which four are known to infect humans. Dabie bandavirus, severe fever with thrombocytopenia syndrome virus (SFTSV), poses serious threats to public health worldwide. SFTSV is a tick-borne virus mainly reported in China, South Korea, and Japan with a mortality rate of up to 30%. To date, most immunology-related studies focused on the antagonistic role of SFTSV non-structural protein (NSs) in sequestering RIG-I-like-receptors (RLRs)-mediated type I interferon (IFN) induction and type I IFN mediated signaling pathway. It is still elusive whether the interaction of SFTSV and other conserved innate immune responses exists. As of now, no specific vaccines or therapeutics are approved for SFTSV prevention or treatments respectively, in part due to a lack of comprehensive understanding of the molecular interactions occurring between SFTSV and hosts. Hence, it is necessary to fully understand the host-virus interactions including antiviral responses and viral evasion mechanisms. In this review, we highlight the recent progress in understanding the pathogenesis of SFTS and speculate underlying novel mechanisms in response to SFTSV infection.The genus  Bandavirus, classified under the order Bunyavirales, family Phenuiviridae, comprises 7 species, including Dabie bandavirus, Bhanja bandavirus, Guertu bandavirus, Heartland bandavirus, Hunter Island bandavirus, Kismaayo bandavirus, and Lone Star bandavirus, which are represented by Dabie bandavirus (SFTSV), Bhanja virus (BHAV), Guertu virus (GTV), Heartland virus (HRTV), Hunter island virus (HUIV), Kismaayo virus (KISV), and Lone star virus (LSV) respectively.According to the International Committee on Taxonomy of Viruses (ICTV) , the genDabie bandavirus, known as severe fever with thrombocytopenia syndrome virus, is a negative-sense single-strand RNA virus that was first discovered in 2009 and mainly reported in China, South Korea, Japan, and most recently in Vietnam, Thailand, and Pakistan  and occasionally from person to person via contacting the blood of SFTS patients directly , mediating the replication of the viral genome. The M segment encodes glycoprotein (Gn and Gc) precursor that is cleaved into two glycoproteins, Gn and Gc, mediating virion assembly and viral entry into cells. The S segment encodes a non-structural protein (NSs) in the sense orientation and the nucleoprotein (NP) in the antisense orientation, serving as an important virulence factor and mediating genome replication and virion assembly respectively , Medium (M), and Small (S) segments ectively .via recognition of viral RNA directly, such as eponymous retinoic acid-inducible gene 1 (RIG-I), mainly sensing short (<300 base pairs) dsRNA with a 5′triphosphate (5′ppp), which can be formed during SFTSV infection . Nucleic acids generated from viruses are one of the typical pathogen-associated molecular patterns (PAMPs), which could be specifically targeted by different nucleic acid pattern recognition receptors (PRRs). RIG-I-like-receptors (RLRs) are important in sensing RNA virus infections nfection , 21. Actnfection . Accordinfection . SFTSV Nnfection –27, whicnfection , 29. Hervia lysosome membrane invagination, CMA refers to take up substrate proteins with KFERQ-motif selectively via lysosome-associated membrane protein type 2A (LAMP-2A), while autophagy is the best characterized one, involved in engulfing cellular materials into double-membrane vesicles for lysosome degradation. To date, multiple studies have clarified the complicated interconnections of the viral life cycle and the autophagic process , and macroautophagy (hereafter referred to as autophagy). Of which, microautophagy is involved in taking up cytosolic material directly  process , 31. TypBunyavirales affected the host autophagy process and the role of autophagy in Bunyavirales infection. Canonical TLRs are reported to participate autophagy process in macrophages. TLR7 but not other TLRs might interact with and recognize RVFV glycoprotein directly  is an evolutionarily conserved signaling pathway, induced by ER stress, which plays a critical role in sensing and removing the accumulation of misfolded proteins and restoring homeostasis , 40. The, but not other viral proteins  of SFTSV, served as the inducer of the UPR  . Compareexosomes –57. Undeexosomes . mtDNA ivia classical dsRNA sensor melanoma differentiation-associated gene 5 (MDA5) . SUV3 and PNPase deficiency were found to dampen the degradation of mtdsRNA , which can be degraded by mtdsRNA enzyme complexes RNA helicase SUV3 and polynucleotide phosphorylase (PNPase). Recently, mtdsRNA was found serving as a novel DAMP, initiating type I IFN response  mtdsRNA . Also, B mtdsRNA . p53 def mtdsRNA .SFTSV infection disturbed mitochondrial homeostasis, induced mitochondrial damage, and promoted mitochondrial DNA oxidization and cytosolic release . We may via TRIF, while TLR7 and TLR8 utilize the adaptor MyD88 to activate IRF3, IRF5, IRF7, and NF-κB. Of those TLRs in endosomes, TLR3 was recently found capable of recognizing alcohol-associated stress-induced mtdsRNA accumulation , including RNA sensor TLR3, TLR7, and TLR8, are located in endosomes, of which TLR3 senses dsRNA, whereas TLR7 and TLR8 are activated by ssRNA. Activated TLR3 mediates the activation of IRF3 and NF-κB mulation . TLR3 exmulation . Howevermulation . Hence, pro-Il-1β and inflammasome-associated components. Activated NLRP3 protein then recruits adaptor protein ASC and pro-caspase-1, inducing the pro-caspase-1 autoproteolytic cleavage into the active form caspase-1, promoting the maturation and secretion of pro-inflammatory cytokines IL-1β and IL-18 and driving gasdermin D (GSDMD)-dependent pyroptosis , TLR9, NLRC4, and AIM2 , 81. cGA366 and Leu374 is important for the activation of STING  was identified as a novel nuclear dsRNA sensor (previously known as nuclear matrix protein hnRNPU) for both DNA and RNA viruses. Like hnRNPA2B1, SAFA was also a nucleic acid binding protein belonging to the hnRNP family. Upon sensing viral RNA in the nucleus, nuclear matrix protein SAFA oligomerized and promoted antiviral gene expression . MyeloidThe induction of SAFA by the virus was severely impaired in ELF4, STING, or MAVS deficient cells. Further studies indicated that ELF4 significantly promoted the expression of SAFA and the phenomena were further enhanced by STING and MAVS overexpression, indicating STING and MAVS may promote the expression of SAFA. The authors hypothesized that SAFA may take function through the STING or MAVS-dependent antiviral IFN pathway and was required for IRF3 and IRF7 recruitment to the IFN-β promoter. Based on the current information, SAFA took function in a cytoplasm-independent manner under VSV or HSV-1 infection, which cannot affect the activation and phosphorylation of IRF3. The detailed function of SAFA under virus infection is largely elusive. It will be an interesting topic to investigate whether SAFA can be translocated to cytoplasm like hnRNPA2B1 under infection status.via death receptors, such as TNF receptor, Fas, and TRAIL receptor. The intrinsic apoptotic pathway is activated by various intracellular stimuli, such as cytochrome c DNA damage, oxidative stress, and ER stress. Of note, mitochondria are crucial for the activation of apoptosis  taking part in the immune escape of those innate immune responses? How can SFTSV hijack the host process for viral replication? How can autophagy participate in sensing and defending SFTSV challenges? Is the UPR related to SFTSV-induced autophagy and inflammation? What are the detailed mechanisms of SFTSV in inducing mitochondrial stress? Are mitochondria playing a key role in host defense against SFTSV infections? Are mtdsRNA and mtDNA taking part in the modulation of classical RNA and DNA PRRs under the SFTSV challenge? Are the nucleic acid sensors such as SAFA or hnRNPA2B1 capable of sensing SFTSV infection? If so, what is the immune escape mechanism of SFTSV? Answering and discovering those possible interconnections will not only enhance our understanding of the viral mechanism to interact with host immunity but also lead to the potential anti-SFTSV strategies.Conceptualization, C-mZ. Original draft preparation, C-mZ. Writing—review and editing, C-mZ and X-jY. Project administration, C-mZ and X-jY. Funding acquisition, C-mZ and X-jY. All authors contributed to the article and approved the submitted version.This study was supported by the National Natural Science Funds of China (81971939 and 31570167) and the Fundamental Research Funds for the Central Universities (2042021kf0046). The funders had no role in the study design, data collection and analysis, decision to publish, or the preparation of the manuscript.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


P = 0.009 in adults), southern provinces , and western provinces . The results in this study suggest that cervid species might serve as important hosts for the transmission of Aino virus, highlighting the need for careful monitoring of Aino virus infections in cervids.Aino virus is an infectious, non-contagious, vector-borne agent that has been implicated in arthrogryposis-hydranencephaly syndrome in newborn cattle, sheep, and goats. Information about reservoirs and host animal species susceptible to Aino virus remains unclear. To further explore the role of cervids in Aino virus infection transmission, we investigated cervid sera to determine the prevalence of Aino virus-neutralizing antibodies and to identify factors correlated with antibody positivity. We screened cervid serum samples collected in the Republic of Korea to better understand infection patterns in this animal species. Overall, Aino virus infection was widespread; 75 of 716  farmed-cervid serum samples collected from 292 herds contained antibodies to Aino virus. Serological evidence of Aino virus infection was demonstrated in 5 of 43 free-ranging cervids, accounting for a prevalence rate of ~11.6% (95% CI = 4.6–26.4). Our results revealed that age class and geographic location affected seroprevalence. The main risk factors associated with Aino virus seroprevalence were older age (> 2 years old, OR = 2.221, 95% CI = 1.209–4.079,  Peribunyaviridae, genus Orthobunyavirus. Aino virus is an infectious, non-contagious, vector-borne agent that has been implicated in arthrogryposis-hydranencephaly syndrome in calves  were 4.5 and 13.3%, respectively  based on methods described by Humphry et al. . A totalHydropotes inermis argyropus) and Siberian roe deer (Capreolus pygargus)] rescued by local rescue parties or housed at captured wildlife rescue centers were available for testing. Cervid blood collection and serum preparation were performed according to previously reported methods  of the Aino virus strain KSA9910  was added to a volume of 50 μL in the test wells of a flat-bottomed microtiter plate and mixed with an equal volume of serum that had been serially diluted in tissue culture medium. After this initial incubation, the virus–serum sample dilutions were dispensed into 96-well microplates containing a monolayer of Vero cells and cultivated as previously described  was carried out to confirm serological evidence of Aino virus infection. Samples were heated to 56°C for 30 min to inactivate complement factors. Vero cells  were maintained in alpha-minimum essential medium  containing 5% fetal bovine serum and antimycotic antibiotics (Gibco). SNTs against Aino virus were performed in flat-bottomed 96-well plates. A 100 50% tissue culture infective dose  the Korea Animal Health Integrated System ; (2) veterinarians in wildlife rescue centers; and (3) animal owners using a questionnaire form according to the methods previously reported . We inveP < 0.05  was considered statistically significant. Statistical analyses in this study were performed by using the using commercially available statistical software .The apparent prevalence and 95% confidence interval (CI)  of true A total of 759 cervids were investigated in this study. Overall, Aino virus infection was widespread, as 75 of 716  farmed cervid serum samples collected from 292 herds were shown to have antibodies to Aino virus as determined by SNTs. In addition, 43 herds analyzed in this study included seropositive animals , as demonstrated in Hydropotes inermis argyropus) and Siberian roe deer (Capreolus pygargus), are excluded from Serological evidence of Aino virus infection was found in 5 of 43 free-ranging cervids that were rescued by local rescue parties or were housed at captured wildlife rescue centers, accounting for a serological prevalence of ~11.6% (95% CI = 4.6–26.4) , as deteP = 0.009 in adults). The high seroprevalence in adult animals likely represents continuous episodes of transmission. Additionally, there were substantial regional differences in seroprevalence within the ROK. We observed a significant difference in the individual likelihood of positivity between southern provinces and northern provinces . We also observed a significant difference in the individual likelihood of positivity between western provinces and eastern provinces . Differences in seroprevalence between herd size and species were not significant (P > 0.05). All the Aino virus-SNT-positive cervid serum samples in this study were found to be serologically negative and did not cross-react with Akabane virus, which belongs to the Simbu serogroup of the genus Orthobunyavirus.Our cervid serum sample analysis results collected in 2011 revealed that age class and geographic location affected seroprevalence. In the univariable analyses , older aAino virus seroprevalence was estimated to be 11.8% (farmed cervids 10.5% and free-ranging cervids 14.7%), demonstrating that exposure to this virus is prevalent among farmed and free-ranging cervids in the ROK despite the paucity of reported outbreaks, probably due to often unapparent clinical signs and underascertainment of Aino virus data.Orthobunyavirus, similar to Akabane virus and Aino virus. Although Peaton virus and Shamonda virus were isolated sequentially in Japan, no Peaton or Shamonda virus infection cases have been reported in the ROK. Only one paper published in 2018 demonstrated serological evidence of Peaton and Shamonda viruses in a small number of animals , without any clinical signs. Additionally, no cross-reactivity among Akabane virus, Aino virus, Peaton virus, and Shamonda virus was observed in neutralization tests  , native The current study represents the first assessment of factors associated with Aino virus seropositivity in cervids in the ROK. The seroprevalence of Aino virus in cervids was significantly different among age cohorts and geographical locations. Aino virus prevalence among cervids in this study was highest in the older group, which may represent continuous transmission. Our results also demonstrated that the prevalence of Aino virus antibodies varied widely in the ROK. The reasons for the region-specific differences in Aino virus transmission are unclear, but the prevalence of Aino virus was higher in the southern and western regions of the ROK. One possibility is that there may be ecological and climatic factors that promote increased exposure to infected reservoirs in tropical rainforest regions. Alternatively, viral persistence or transmissibility may be higher in regions with elevated temperatures and rainfall.The results in this study demonstrated that Aino virus infection was prevalent among the farmed and free-ranging cervids surveyed; approximately one in 10 animals were infected with the virus. The present study demonstrates that Aino virus is widely distributed in the ROK and that although an outbreak of Aino virus among native cervids in the ROK has yet to be reported, susceptible cervids are at risk of becoming infected with Aino virus. The results in the present study suggest that cervid species might serve as important hosts for the transmission of Aino virus, highlighting the need for closer monitoring of Aino virus infections in cervids in the ROK. The results of this seroprevalence study may serve as a basis for future epidemiological studies of Aino virus infection in the ROK.The original contributions presented in the study are included in the article/supplementary material, further inquiries can be directed to the corresponding author/s.J-YY and YJG conducted the laboratory experiments and wrote the first draft. J-YY conceived and designed the study and revised the manuscript. All authors contributed to manuscript revision, read the manuscript, and approved the submitted version.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


We propose a theoretical model to elucidate intermolecular electrostatic interactions between a virus and a substrate. Our model treats the virus as a homogeneous particle having surface charge and the polymer fiber of the respirator as a charged plane. Electric potentials surrounding the virus and fiber are influenced by the surface charge distribution of the virus. We use Poisson–Boltzmann equations to calculate electric potentials. Then, Derjaguin’s approximation and a linear superposition of the potential function are extended to determine the electrostatic force. In this work, we apply this model for coronavirus or SARS-CoV-2 case and numerical results quantitatively agree with prior simulation. We find that the influence of fiber’s potential on the surface charge of the virus is important and is considered in interaction calculations to obtain better accuracy. The electrostatic interaction significantly decays with increasing separation distance, and this curve becomes steeper when adding more salt. Although the interaction force increases with heating, one can observe the repulsive–attractive transition when the environment is acidic. I.Intermolecular force is important to determine the stability and persistence of viruses on the surface substrate. Several approaches based on experiment have been performed to elucidate the interaction of respiratory viruses, such as SARS-CoV-2,et al.et al.et al.et al.et al.Several theoretical models have been proposed to study the electrostatic properties of viruses. The soft colloid model of OhshimaSARS-CoV-2 virus has caused an outbreak, resulting in COVID-19 pandemic with many severe consequences.In this work, using SARS-CoV-2 as a case to study, we propose a simple theoretical approach for calculating the interaction of the virus with the surface charge of the polymer fiber of the mask. We employ Poisson–Boltzmann equations to obtain electrostatic potential profiles surrounding objects and combine these potentials with Derjaguin’s approximation to determine interaction forces. Since the surface charge density of particle-like viruses is affected by the local electrostatic field induced by fiber plane, the effects of the induced electric field on our calculations are considered. We also discuss influences of other environmental factors, such as pH, salt concentration, and temperature, on the interaction force to further verify our model.II.In this section, we construct a theoretical description and parameters for the SARS-CoV-2 virus immersed in the electrolyte and polymer fiber of the mask. A.e is the elementary charge and the sum takes over all types of AAs with ± corresponding to deprotonated  and protonated  AAs, respectively. Reff = 70 nm, which is the distance from the center of the virus to S proteins. Because of the screening of the other charge, half of S protein charge N number of S proteins,A SAR-CoV-2 virus consists of a nucleocapsid enveloped by a spherical lipid membrane with S proteins attached on its surface.kB is the Boltzmann constant, T is the ambient temperature, ψ is the potential of the isolated virus, r is the radial distance to the center of a virus, ε0 is the vacuum permittivity, εr = 78 is the permittivity of the electrolyte, εp = 4 is the permittivity of the nucleocapsid, z0 and n are the ionic valence and ion number concentration in solution, respectively. In this work, we consider a monovalent electrolyte, and thus, z0 = 1. In addition, the authors of Ref. κReff ≫ 1, an analytical expression of potential distribution around a spherical particle is approximately obtained byψ0 is the surface potential of the virus, and the relation between surface charge and surface potential isThe electric potential is analytically described by the Poisson–Boltzmann equations associated with the corresponding boundary conditions for a particle-like virus.B.σS = −1 e/nm2. The gradient of electrostatic potential near the fiber obeys the Poisson–Boltzmann equationz = 0 at the surface (as depicted in ϕ(z) plays a role as local potential, which can alter the surface charge distribution of the virus. This effect will be discussed further in Sec. ϕ0 being the surface potential of the plane). An analytical expression of ϕ0 isSince the dimension of the polymer fiber is much larger than the radius of the virus,III.In this section, we use Derjaguin’s approximation and a linear superposition of potentials to calculate the interaction force between the SARS-CoV-2 virus and the electret fiber of the mask. The effect of electret fiber potential on the virus charge surface is considered. We also delve into how the calculated force depends on the separation distance and environmental factors, including pH and ion concentration.A.κReff ≫ 1, the interaction electrostatic energy per unit area of this part and planar surface is equal to the energy of interaction per unit area of two infinite planes at the same separation distance h [Fpl is the interaction force between the planar part of the spherical particle and substrate per unit area and H is the distance from the virus surface to the fiber. To calculate the virus–fiber interaction, the explicit form of Fpl must be known.To calculate the interaction force, a spherical object is modeled as a superposition of infinitesimal parallel parts as shown in stance h . Thus, tφ being the potential between two planes.When the charge body is immersed in an electrolyte, it exhibits the excess osmotic pressure ΔΠ that originated from the electrical double layer and Maxwell forces. These forces are strongly dependent on the interactions of surface charge body and counterions in the electrolyte. Thus, the total interaction force of two planes can be obtained from the integration of the total stress tensor, which consists of the Maxwell electrostatic and Van’t Hoff osmotic component, over the surface enclosing either one of the two planes Σ,z = ∞ and φ(z) is the linear superposition of potentials of two isolated planes. In general, the surface charge of the spherical virus can be influenced by the planar potential. Therefore, the value of γsp of the plane divided from the spherical particle depends on the separation distance h. Based on Eq. In our calculations, the closed surface is chosen according to the theory of Verwey and Overbeek.From Eqs. B.pH, the surface of the particle-like virus is kept constant. By using Eq. N = 60 S proteins on its surface and the fiber as a function of pH at a salt concentration of 2 mM separation distance of 2.1 nm, and at T = 25 °C is calculated and shown in pH level in this work ranging from 5.6 to 8.4 is consistent with the regular pH level of the respiratory droplet. In this approach, the force is reversed from attractive to repulsive at the isoelectric point of the charge surface with a pH value of 5.6. These results suggest that when a respiratory droplet is attached to the mask with negative charge fiber, the virus within it is more readily to adhere to the substrate in an acidic environment.First, we assume that when the virus reaches the vicinity of the planar fiber, the surface charge density of the virus is not influenced by the planar electrostatic field. In the absence of the local field induced by the fiber, according to Eq. In C.h from the tagged point to the planar. By taking integration over h in Eq. To improve the accuracy of our calculations, we take into account the effects of the electrostatic field of the polymer fiber on the charge surface density of the particle-like virus. The roles of the induced field become more important when the virus is closer to the fiber. The local electrostatic potential of the fiber causes the shift of pH at the ion concentration of 2 mM separation distance of 2.1 nm, and the temperature of 25 °C. One can observe a quantitative good agreement between our calculations and simulation.pH ≈ 6.1. Compared to the case of the constant virus charge surface, the transition point is shifted toward the basic pH region. Therefore, when the virus approaches the mask, under the local electrostatic of the electret fiber, not only the charge distribution but also the overall charge surface is modified depending on the fiber potential. pH. It is reasonable since the potential of the particle-like virus and the plane gradually approach zero at a large distance as one can see from Eqs. N = 60, pH = 7, and T = 25 °C can be observed in n = 100 mM, the force reduces to near zero at H ≈ 5 nm. This value for n = 10 and 2 mM is H ≈ 15 nm and H ≈ 20 nm, respectively. This variation originates from the increase in Debye–Hückel parameter value κ when the salt concentration increases. Upon this condition, the electric double layer thickness around each object decreases, and it leads to the weakening of the screening effect. To further validate our analytical model, we also investigate the dependence of the calculated interaction force on environmental parameters. In In addition to humidity, temperature has a significant effect on the virion in the respiratory droplet. A recent work of Dbouk and DrikakisIV.We have proposed our simple calculation based on the Derjaguin method for the interaction force of the SARS-CoV-2 virus and the electret fiber of the mask. The fiber is treated as an infinite plane while the virus is simplified to a spherical particle with charge of all S proteins smearing uniformly over the particle surface. We also compare our approach with other simple analytical models to unravel the importance of the exact solution of the Poisson–Boltzmann equation as well as the effect of fiber electric potential on the virus charge surface to determine the sign and magnitude of electrostatic interaction. Numerical results calculated by our model show the substantial dependence of interaction force on the surface charge of both object and environmental parameters that include pH, salt concentration, and temperature. The good agreement of these results and the prior work proves the high accuracy of our model. Therefore, with the simple yet precise tool that is established in this work, one can quantitatively calculate the interaction between the virus and the inanimate surface substrates and have better insight into this intermolecular interaction.κReff > 10. For κReff < 5, theoretical predictions can become less accurate.There are two main limitations in our calculations. First, we theoretically treat the SARs-CoV-2 virus as a spherical particle, while S proteins on the surface of the virus make the surface rough. The roughness could lead to the difference between our results and simulation. Second, our model is based on Derjaguin’s approximation, which provides better results when


In the present study, canthaxanthin was produced by biofermentation from Dietzia natronolimnaea HS-1 (D. natronolimnaea) and was loaded in phospholipid vesicles prepared with natural component using an easy and low dissipative method. Indeed, glycerosomes, hyalurosomes, and glycerohyalurosomes were prepared by direct hydration of both phosphatidylcholine and the biotechnological canthaxanthin, avoiding the use of organic solvents. Vesicles were sized from 63 nm to 87 nm and highly negatively charged. They entrapped a high number of the biomolecules and were stable on storage. Canthaxanthin-loaded vesicles incubated with fibroblasts did not affect their viability, proving to be highly biocompatible and capable of inhibiting the death of fibroblasts stressed with hydrogen peroxide. They reduced the nitric oxide expression in macrophages treated with lipopolysaccharides. Moreover, they favoured the cell migration in an in vitro lesion model. Results confirmed the health-promoting potential of canthaxanthin in skin cells, which is potentiated by its suitable loading in phospholipid vesicles, thus suggesting the possible use of these natural bioformulations in both skin protection and regeneration, thanks to the potent antioxidant, anti-inflammatory and antiageing effects of canthaxanthin. Cantharellus cinnabarinus  colorimetric assay. MTT solution  was added to each well, and cells were incubated for 2–3 h. After that, the formed formazan crystals were dissolved in dimethyl sulfoxide, and their concentration was spectrophotometrically quantified at 570 nm by using a microplate reader . Results are shown as percent of cell viability in comparison with nontreated cells (100% viability).Primary mouse embryonic fibroblasts (3T3)  were grown as monolayer in 75 cm2 for 24 h. Cells were stressed with hydrogen peroxide  and treated for 4 h with canthaxanthin in dispersion or loaded in vesicles opportunely diluted to reach 2 mg/mL and 0.2 mg/mL of payload. Cells stressed with hydrogen peroxide and untreated were used as negative control, and healthy cells unstressed and untreated were used as positive control. At the end, the cells were washed with PBS, and the MTT assay was used to assess the viability. Untreated cells were used as 100% of viability, and cells stressed with hydrogen peroxide were used as negative control.The ability of canthaxanthin to protect fibroblasts from damages induced by hydrogen peroxide was evaluated. Cells were seeded in 96-well plates and incubated at 37 °C in 5% CO0) and at 24 h and 48 h (at) to measure the areas during the treatment. The captured images were quantified by Java’s image J software (http://rsb.info.nih.gov (accessed on 2 January 2022)) by measuring the area of the wound The migration of cells towards the wounds was expressed as percentage of wound closure:The ability of canthaxanthin to stimulate cell proliferation and migration was evaluated measuring the speed rate of the wound healing as a function of the time of exposition with the samples (scratch assay). Cells were cultured in 6-well plates until the complete confluence was reached. Then, a linear scratch was generated with a sterile plastic pipette tip. The cells were gently washed with fresh medium to remove the scattered fragments and then treated for 48 h with canthaxanthin in dispersion or loaded in vesicles, properly diluted with medium to reach 2 µg/mL and 0.2 µg/mL of payload. Untreated cells were used as positive control. The changes of the area of the lesion were monitored by using an optical microscope (10 × objective), and the related images were captured at initial time to measure the wounded area immediately after scratching  was used for multiple comparisons of means, and Tukey’s test and Student’s 40H22O2) as detected at m/z 565.4, Canthaxanthin was obtained by biofermentation using D. natronolimnaea HS-1 and an enzymatic hydrolysed molasses as substrate. The concentration of the obtained biomass in the grown medium was ~1.81 mg/L and contained ~1.01 mg/L of canthaxanthin, which was statistical equal to the concentration of biomass, indicating that canthaxanthin was the main carotenoid produced. The identification and quantification of carotenoid pigments contained in the biomass were carried out using the HPLC and the atmospheric pressure chemical ionisation tandem mass spectrometric analysis. The peak reported in Phaffia rhodozyma; indeed its production was lowered when the glucose content increased. This fact might be due to the long-time exposure of dissolved carotenoids to oxygen, light, and microorganisms in the environment leading to an increased chance of oxidation or degradation of the dissolved carotenoids.As previously reported, the concentration of canthaxanthin produced depended on the used concentration and kind of substrate, and using molasses increased when its concentration increased from 5 g/L to 25 g/L ,23. On tA preformulation study was carried out to select the most homogenous dispersions containing stable and small vesicles. Different kinds and numbers of phospholipids and various water co-solvents and/or biopolymers were used, and their ability to incorporate an increasing amount of canthaxanthin was evaluated as well. Among all, soy lecithin (120 mg/mL) was selected as the phospholipid due to its direct plant origin (without transformation process) and the suitable characteristics of formed vesicles. A total of 20 mg/mL of canthaxanthin was selected as the most suitable loading concentration. Glycerol (30%) was used to partially substitute the water, and hyaluronan (0.1%) was also added to improve the homogeneity and stability of the resulting vesicles. The average diameter, polydispersity index, and surface charge of the vesicles were measured and compared as a function of the used components, using empty vesicles as reference .p > 0.05 among the size of samples), were monodispersed (polydispersity index ~0.24), and highly negatively charged (~55 mV). The incorporation of canthaxanthin into liposomes and glycerosomes caused a slight increase in the mean diameter of the vesicle, which was ~71 nm (p > 0.05 between the mean diameter of two samples and p < 0.05 versus the mean diameter of other empty and loaded vesicles). The incorporation of canthaxanthin into hyalurosomes addressed a further increase in mean diameter up to ~87 nm (p < 0.05 versus the mean diameter of other vesicles). The loading effect was negligible in glycerohyalurosomes with their size being similar to that of empty vesicles . The vesicle enlargement may be related to the intercalation of canthaxanthin inside the bilayer, which allowed a slightly modification of phospholipid assembling. The simultaneous presence of glycerol and hyaluronan deleted this effect. The surface charge of all the canthaxanthin-loaded vesicles was ~−50 mV, similar to that of the corresponding empty vesicles, indicating that it was mostly intercalated inside the bilayer and not located in the vesicle surface. The amount of canthaxanthin effectively incorporated in glycerosomes, hyalurosomes, and glycerohyalurosomes was higher  than that in liposomes (~57%), confirming that glycerol and sodium hyaluronate favoured the loading of canthaxanthin.Empty vesicles were the smallest irrespective of the used components  was 51% ± 1 and slightly increased up to 58% ± 1 after its loading into the vesicles. The result underlined the suitability of prepared vesicles; indeed, they did not negatively affect the antioxidant activity of canthaxanthin, on the contrary, they provided a synergic effect due to the antioxidant molecules naturally occurring in lecithin ,23.p > 0.05 among the values at different dilutions). The treatment of the inflamed cells with canthaxanthin loaded into vesicles inhibited the nitric oxide release in a dose- and composition-dependent manner. Indeed, any inhibition  was detected by using the higher dilution (concentration 0.02 μg/mL), except when liposomes were used, as the inhibition was ~40% (p < 0.05 versus others). When canthaxanthin was loaded in glycerosomes, hyalurosomes, and glycerohyalurosomes at lower dilutions , the nitric oxide production was almost completely inhibited , except using glycerosomes at an intermediate dilution (concentration 0.2 μg/mL), which was not effective.Nitric oxide is expressed in large amounts in cells stimulated with lipopolysaccharide. This overexpression has been demonstrated to be associated in the pathogenesis of acute and chronic inflammatory conditions . The capp > 0.05 among the values obtained with different dilutions). The incubation with the canthaxanthin-loaded liposomes achieved similar values . The incubation with canthaxanthin loaded in glycerosomes, hyalurosomes, and glycerohyalurosomes increased the cell viability up to ~110%, even if the values were not statistically different from that obtained using the dispersion . Thus, the results indicate that the loading of canthaxanthin in these vesicles can address a proliferative effect in fibroblasts [The most representative cells of the dermis are fibroblasts (3T3), which are mainly involved in skin repair. Given that, they were used to evaluate in vitro the potential cytotoxicity of samples . The celroblasts ,25,27.p < 0.05 versus values obtained using the vesicles) . The treatment with canthaxanthin loaded in vesicles ensured a complete protection of fibroblasts, as the cell viability was ≥100%. The protection was formulation depending as the viability of cells treated with canthaxanthin (2 μg/mL) loaded in liposomes and glycerosomes was ~100% , and that of cells treated with 0.2 μg/mL of canthaxanthin loaded in glycerosomes, hyalurosomes, and glycerohyalurosomes was ~120% . The loading improved the effectiveness of the payload probably favouring a better internalisation of it inside the cells and their membrane, which is widely damaged by reactive oxygen species [Hydrogen peroxide can decrease the viability of human skin fibroblasts with a concurrent increase in generation of reactive oxygen species and cell apoptosis . Many stesicles) . The tre species .An in vitro scratch assay was carried out on a cell monolayer of fibroblasts to evaluate the capability of canthaxanthin in dispersion or loaded in vesicles of favouring the wound closure promoting cell proliferation and migration . The untD. natronolimnaea and molasses as the inexpensive and renewable substrate in place of glucose, and it was loaded in natural-based vesicles prepared with soy lecithin, glycerol, hyaluronan, and water. The applied biotechnological methodologies addressed the microbial production of canthaxanthin in high yield by the valorisation of an agrifood byproduct and avoided the new glucose dissipation, as confirmed in previous studies [D. natronolimnaea culture process and can also be used as a guideline for similar microbial cultivation systems. The biotechnological production of canthaxanthin has been combined with the formulation of nanotechnological carriers, which previously revealed to be optimal carriers for skin delivery of natural molecules and carotenoids [There is an increasing demand for carotenoids, such as canthaxanthin, due to their applications in the pharmaceutical, cosmeceutical, flavour food, and feed industries; however, the extraction and synthesis of these compounds can be expensive, technically challenging, and environmentally hazardous. The biotechnological production of canthaxanthin may provide an attractive alternative at the industrial level, which meets the growing demand to reduce costs and environmental pollution and avoid geographical and seasonal limitations ,32. An a studies . Accordi studies  previousotenoids . They arotenoids . They weotenoids . Liposomotenoids . The obtotenoids . Canthaxotenoids ,35,36,39otenoids ,40. HoweThe active interaction of phospholipid vesicles with the cells and their ability to favour payload internalisation, has been confirmed in previous studies. According to this, the treatment of a scratched area performed in a cell monolayer, with canthaxanthin-loaded vesicles, promoted the proliferation and migration of fibroblasts thus speeding up the closure of the wounded area in comparison with the aqueous dispersion of canthaxanthin. In particular, the combination of glycerol and hyaluronan seems to be the most promising as the complete closure during 48 h was detected only treating the cells with this formulation.D. natronolimnaea culture and molasses was confirmed as well as its efficacy in the skin when loaded into phospholipid vesicles, especially hyalurosomes and glycerohyalurosomes, which appeared as the most promising system for the treatment of skin damages.The demand of natural and safe products is increasing exponentially nowadays due to the awareness that a less polluted world ensures a better lifestyle. Even the synthesis of new molecules has been reduced in favour of the biotechnological production of both innovative and well-known bioactive molecules. In the present study, the effective and scalable bioproduction of canthaxanthin using


None of the 180 seedlings originating from TYLCSV-infected plants showed any phenotypic alteration typical of virus infection. Moreover, whole viral genomic molecules could not be detected in their cotyledons and true leaves, neither by membrane hybridization nor by rolling-circle amplification followed by PCR, indicating that TYLCSV is not a seed-transmissible pathogen for tomato. Examining the localization of TYLCSV DNA in progenitor plants, we detected the virus genome by PCR in all vegetative and reproductive tissues, but viral genomic and replicative forms were found only in leaves, flowers and fruit flesh, not in seeds and embryos. Closer investigations allowed us to discover for the first time that these embryos were superficially contaminated by TYLCSV DNA but whole genomic molecules were not detectable. Therefore, the inability of TYLCSV genomic molecules to colonize tomato embryos during infection justifies the lack of seed transmissibility observed in this host.Seed transmission is an important factor in the epidemiology of plant pathogens. Geminiviruses are serious pests spread in tropical and subtropical regions. They are transmitted by hemipteran insects, but a few cases of transmission through seeds were recently reported. Here, we investigated the tomato seed transmissibility of the begomovirus  Geminiviridae family, with nine accepted genera and an increasing number of unassigned species, is the largest family of plant-infecting viruses [Begomovirus genus is the largest genus of plant viruses and includes more than 400 assigned species [The  viruses . Their s viruses . Within  species ; it coun species . sweet potato leaf curl virus [tomato yellow leaf curl virus (TYLCV) [tomato leaf curl New Delhi virus [mung bean yellow mosaic virus [bitter gourd yellow mosaic virus [pepper yellow leaf curl Indonesia virus [Bemisia tabaci [Until now, geminiviruses have been commonly believed to be naturally transmitted by hemipteran insects, such as whiteflies, leafhoppers, plant-hoppers and aphids. Recently, seed transmissibility of some begomoviruses, such as rl virus , tomato  (TYLCV) , tomato hi virus ,7, mung ic virus , bitter ic virus  and peppia virus  has beenia virus ,11 or toa tabaci . Howevera tabaci ,13,14,15Tomato yellow leaf curl Sardinia virus (TYLCSV), another monopartite begomovirus, is present in the entire Mediterranean basin [an basin  and, in an basin . ClarifyAgrobacterium tumefaciens LBA4404 cultures harboring a 1.8mer TYLCSV construct (Genbank Acc. No. X61153) [0; 1). Seeds collected from eight infected plants were surface-sterilized for 10 min in 70% ethanol, then left for 10 min in 10-fold diluted commercial bleach, followed by several washes in sterile water before sowing. For analysis on G1 seedlings, 20 to 24 seeds from each G0 plant were used. G1 plants were maintained in an insect-free greenhouse and monitored for the appearance of symptoms during their life cycle (Tomato plants (cv. Moneymaker) were inoculated at the four-leaf stage by  X61153) , while afe cycle A. CotyleTotal DNA was extracted by the dot-blot method  from all2 and 1 unit per µL Platinum Taq DNA polymerase . The mixture was denatured for 4 min at 95 °C, followed by 35 cycles of 30 s at 95 °C, 30 s at 58 °C and 30 s at 72 °C and by a final cycle at 72 °C for 10 min. The mixture was loaded onto 1% agarose gels, and the gels were run at 100 V, stained with ethidium bromide and visualized under UV light.PCR assays were carried out in 25 µL reactions containing 1× PCR buffer, 200 µM each of dNTPs, 0.4 µM of primers , 2 mM MgQuantitative real-time PCR (qPCR) was carried out using iCycler iQTM Real-Time PCR Detection System , as described previously , with thDNA samples (approximately 300–500 ng) were loaded onto 1% agarose gel and separated in 0.5 × TBE, containing 0.5 µg/mL ethidium bromide, electrophoresed at 70 V for 3 h and blotted onto positively charged nylon membranes . Membranes were then hybridized with a digoxygenin-labeled coat-protein-specific probe obtained with the TY1(+)/TY2(−) primers , followi1 material (cotyledons and true leaves) was diluted 1 to 5 in water and subjected to RCA using the TempliPhi Kit , according to the manufacturer’s instructions. A DNA extract from an infected tomato plant was used as positive control, while extracts derived from G1 seedlings obtained from a healthy plant were used as negative controls. RCA products (diluted 1 to 5 in water) were used for end-point PCR amplification.DNA extracted from GTo investigate the seed transmissibility of TYLCSV, tomato plants were agroinoculated with a TYLCSV clone and maintained under insect-free conditions until flower production and fruit maturation A. Mild c0 plants, deriving from two independent inoculation experiments, were used in two separate grow-out assays (1 seedlings (n = 180) were analyzed for their phenotype and for the presence of viral DNA. Overall, none of the plants displayed any symptom that could be ascribed to TYLCSV infection, at least up to two months after sowing. When the presence of viral DNA was investigated in cotyledon and true leaf extracts of G1 seedlings, using a Southern blot developed with a virus-specific probe targeting the coat protein gene, viral genomic forms could not be detected /TY2(−). However, even in this case, no TYLCSV-related amplicons could be visualized . 1 seedlings or, at least, that its amount is below the detection limits of the adopted techniques, suggesting that it is not seed-transmissible in tomato. Taken together, these results indicate that TYLCSV DNA cannot be detected in G0 infected plants, such as petals, sepals, pistils, stamens, fruit flesh, seeds and embryos, and subjected them to end-point PCR analysis using the TY1(+)/TY2(−) primers. Positive signals were obtained from leaves and extracts prepared from all the organs considered   . These i < 0.05) C. 1 seedlings. For this, three new batches of embryos (n = 18–20 each) were collected from infected seeds. Once verified by PCR that TYLCSV DNA could be amplified from all extracts (The reduced concentration of TYLCSV DNA in seeds and embryos detected by qPCR and the inability to detect genomic or replicative forms by a membrane-based approach indicated a poor accumulation, if any, of viral genome in these organs. Therefore, we adopted the RCA-based approach above used to identify circular genomic DNAs in GThis prompted us to verify if the amplification products obtained from embryos by direct PCR, without performing RCA, could result from a contamination of the external layers of the embryos by the surrounding tissues. For this, embryos were subjected to a surface-sterilization step and their DNA extracts were processed by PCR and RCA/PCR, as above. Notably, following such surface treatment, it was possible to successfully amplify the endogenous 25S rRNA gene , while tOverall, these results allow us to conclude that embryos dissected from infected seeds are surface-contaminated by TYLCSV DNA, originating from the surrounding maternal tissue during seed formation or during the processes of embryo dissection and manipulation. The most important result of these analyses is that no circular genomes of TYLCSV can be detected in embryos, neither in the external layers, nor in the inner tissues of such seed portions, implying that whole viral molecules do not reach embryos, in line with the observed lack of TYLCSV transmission through seeds. Ty genes-based resistance [Considering the importance of TYLCD and the continuous intercontinental spread of its vector to new areas , biologisistance ,29 and isistance ,31. 1 seedlings. These results are clearly in contrast with the 70 to 85% rates of seed transmissibility reported for TYLCV in tomato [Nicotiana benthamiana, a laboratory solanaceous host highly susceptible to TYLCV and supporting high viral accumulation [1 seedlings reported for TYLCV DNA in tomato [−94, based on the binomial distribution calculation. This probability increases to 10−18 and 10−8 if we consider the lower infection rates of G1 seedlings reported for the same virus in other hosts, such as soybean and pepper, with infection rates of 21% [Our grow-out assays showed not only that the progeny of infected plants did not display viral symptoms, but also that TYLCSV DNA and, more importantly, whole genomic viral molecules are undetectable in the cotyledons and true leaves of Gn tomato ,14 and imulation . It is ws of 21%  and 10% s of 21% , respects of 21% . TherefoIt is worth mentioning that the experiments here reported were conducted with plants artificially inoculated with an infectious clone of TYLCSV , throughSeed transmission depends on the ability of a virus to reach the reproductive organs of the host, facing the problem of crossing physical barriers of the different organs, invading different kinds of tissues and overcoming the physiological alterations that occur during seed maturation and storage. To elucidate such biological process and gain proof of the efficient localization of the pathogen in reproductive structures, it is important, not only to amplify portions of viral genomic sequences, but also to focus on the presence of whole viral genomes using suitable molecular techniques. In this work, we initially verified that TYLCSV does reach the reproductive organs of infected tomato plants, including flowers, fruits and seeds, and that, similarly to TYLCV , its titPrevious reports of geminivirus seed transmissibility often relied on the successful PCR amplification rates of the viral genome, even in the absence of symptoms ,6,7,8,34Given the controversial results reported for TYLCV in tomato ,12,13,14


High Plains wheat mosaic virus. Hence, High Plains virus, maize red stripe virus, wheat mosaic virus, and High Plains wheat mosaic virus (HPWMoV) are synonyms for the causal agent of High Plains disease.In 1993, severe mosaic and necrosis symptoms were observed on corn (maize) and wheat from several Great Plains states of the USA. Based on the geographical location of infections, the disease was named High Plains disease and the causal agent was tentatively named High Plains virus. Subsequently, researchers renamed this virus as maize red stripe virus and wheat mosaic virus to represent the host and symptom phenotype of the virus. After sequencing the genome of the pathogen, the causal agent of High Plains disease was officially named as High Plains wheat mosaic virus is one of the 21 definitive species in the genus Emaravirus in the family Fimoviridae.The genomic RNAs are encapsidated in thread‐like nucleocapsids in double‐membrane 80–200 nm spherical or ovoid virions.The HPWMoV genome consists of eight single‐stranded negative‐sense RNA segments encoding a single open reading frame (ORF) in each genomic RNA segment. RNA 1 is 6,981‐nucleotide (nt) long, coding for a 2,272 amino acid protein of RNA‐dependent RNA polymerase. RNA 2 is 2,211‐nt long and codes for a 667 amino acid glycoprotein precursor. RNA 3 has two variants of 1,439‐ and 1,441‐nt length that code for 286 and 289 amino acid nucleocapsid proteins, respectively. RNA 4 is 1,682‐nt long, coding for a 364 amino acid protein. RNA 5 and RNA 6 are 1,715‐ and 1,752‐nt long, respectively, and code for 478 and 492 amino acid proteins, respectively. RNA 7 and RNA 8 are 1,434‐ and 1,339‐nt long, code for 305 and 176 amino acid proteins, respectively.HPWMoV can infect wheat, corn (maize), barley, rye brome, oat, rye, green foxtail, yellow foxtail, and foxtail barley. HPWMoV is transmitted by the wheat curl mite and through corn seed.Genetic resistance against HPWMoV in wheat is not available, but most commercial corn hybrids are resistant while sweet corn varieties remain susceptible. Even though corn hybrids are resistant to virus, it still serves as a green bridge host that enables mites to carry the virus from corn to new crop wheat in the autumn. The main management strategy for High Plains disease in wheat relies on the management of green bridge hosts. Cultural practices such as avoiding early planting can be used to avoid mite buildup and virus infections. Emaravirus in the family Fimoviridae.HPWMoV, an octapartite single‐stranded RNA virus of negative polarity, is the causal agent of High Plains disease of wheat and corn belongs to the genus  Because the wheat curl mite transmits all the members of the WSMD complex, wheat coinfected by two or three viruses is common in growers’ fields with exacerbated disease phenotype with occasional plant death  coding for the 32 kDa NC protein , QGXXXXXSS1200‐1208 (motif B), SDD1241‐1243 (motif C), KK1284‐1285 (motif D), and EFLST1294‐1298 (motif E)  encodes a 266 kDa protein of RNA‐dependent RNA polymerase  comprising 2,272 amino acids with 36%–42% sequence identity with other reported emaraviral RdRp proteins  and 3B , with 12.5% sequence divergence Figure . RNA 3A RNA 4 1,682 nt) encodes a polypeptide of 42 kDa (P4) with 364 amino acids  and RNA 8  encode 36 kDa (P7) and 21 kDa (P8) proteins with 305 and 176 amino acids, respectively  with that of NE isolate with seven indels in the 5′‐NTR Stewart, . The P5,66.1HPWMoV is transmitted by the wheat curl mite , rye brome , oat (Avena sativa), and rye . Recently, Abdullah et al. (Setaria glauca) and foxtail barley (Hordeum jubatum) plants in Canada.HPWMoV was originally reported as infecting wheat and corn  ratio, indicating a single dominant gene is responsible for resistance in B73. The resistance allele in B73 was linked to marker bn16.29 on the short arm of chromosome 6.Because HP disease was first observed with more symptom severity in corn compared to wheat, Marçon et al.  screenedWsm1, Wsm2, and Wsm3 genes, respectively  genes, Cmc1/Cmc4, Cmc2, or Cmc3, with DNA transferred from Aegilops tauschii, Agropyron elongatum, and S. cereale, respectively, have been reported to provide genetic resistance against wheat curl mite  resulted in adaptation of resistance by wheat curl mite populations to the Cmc gene, enabling them to readily colonize the wheat (Harvey et al., Because HPWMoV is transmitted through wheat curl mites, the development of crop plants resistant to wheat curl mites could provide an effective management strategy not only to HP disease but also against WSMV and TriMV. Additionally, the development of mite‐resistant crop plants could prevent or minimize losses incurred due to wheat curl mite feeding. Thus, mite‐resistant crop plants will provide dual protection from viruses as well as feeding damage by wheat curl mites. So far, in wheat, four curl mite colonization (9.3Control of wheat curl mites through pesticide application for management of HP disease is ineffective due to the secluded nature of mites within the whorl of wheat plants. Because HPWMoV most often occurs in mixed infections in wheat (Burrows et al., For winter wheat, avoiding early planting is recommended to reduce the overlap of green bridge hosts and the new crop wheat. This reduces the chances of direct infestation from the green bridge host, but it also reduces mite and virus buildup and spread during favourable warm autumn temperatures. The effective management of HP disease should include the integration of cultural practices and genetic resistance. Thus, considerably more needs to be learned about HPWMoV resistance relationships in developing wheat lines with WSMV resistance.In areas where winter wheat is grown in cropping systems that include corn, management of HPWMoV in commercial corn is largely accomplished through virus resistance in corn hybrids. This eliminates the impact of the virus on corn production. Corn can still serve as a green bridge host to carry mites and virus to surrounding wheat fields Knoell, , so it i10High Plains wheat mosaic virus in the genus Emaravirus, family Fimoviridae (Tatineni et al., Emaravirus that contains eight genomic RNA segments. Subsequent sequencing of several isolates from wheat, corn, and barley from Kansas and Ohio revealed sequence diversity with two RNA 3 segments in isolates GG1 and KS7 but only one RNA 3 segment in Ohio wheat isolates (Stewart, The causal agent of HP disease was determined to be an octapartite negative‐sense RNA virus, and it was named Stewart, . The seqEmaravirus (Verchot et al., The next step after sequencing the HPWMoV genomic RNAs is determining the functions of virus‐encoded proteins. Based on sequence similarity, the genes encoded by RNA 1, 2, 3, and 4 of HPWMoV were identified as replication‐associated proteins, glycoprotein‐precursor, nucleocapsid, and movement protein, respectively (Tatineni et al., The development of sensitive and broad‐spectrum detection methods will facilitate the detection of HPWMoV isolates in alternate hosts. Though PCR‐based methods were reported (Arif et al., Viruses are obligate parasites, hence they must depend on host and vector machinery for replication, movement, disease development, and transmission. Examination of virus–host interactions will facilitate the identification of host factors that can be used as targets to disrupt the virus life cycle by targeting host proteins. Identification of host factors that do not have a crucial role in plant growth and development but are required for virus life cycle could be candidates for use in the management of viral diseases. The mechanisms of wheat curl mite transmission of HPWMoV are not known. It is not known whether wheat curl mites transmit the virus in a persistent, nonpersistent, or semipersistent manner or if the virus replicates within the mite. Virus–vector interaction studies will facilitate the identification of wheat curl mite proteins required for HPWMoV transmission. These mite proteins could be used as targets to knockout wheat curl mite transmission of HPWMoV. Studies on HPWMoV gene functions and identification of host and vector proteins interacting with HPWMoV proteins could eventually facilitate new management strategies. Candidate host proteins involved in disease development could be targeted via CRISPR/Cas9 technology to interrupt virus–host interactions and mitigate the viruses’ ability to infect plants.Seed transmission of HPWMoV in corn creates serious issues in sweet corn because of the susceptibility of sweet corn hybrids. This is also an issue in hybrid seed corn production that is targeted for export as seed lots testing positive for HPWMoV are at risk of embargo. Understanding the virus–host relationship that leads to HPWMoV seed transmission could enable the development of technology targeting the interruption of seed transmission.Wsm genes provide resistance against WSMV. However, it is not known whether these cultivars provide similar resistance against HPWMoV. It will be important to examine whether Wsm‐based corn and wheat cultivars provide a similar level of resistance to HPWMoV. If any of these genes are also resistant to HPWMoV in wheat, gene pyramiding of these genes with wheat curl mite resistant Cmc genes could provide a dual resistance to virus and vector.Corn and wheat cultivars with The authors declare no conflict of interest.


The economic burden of stroke on the healthcare system has been previously described, but sex differences in healthcare costs have not been well characterized. We described the direct person-level healthcare cost in men and women as well as the various health settings in which costs were incurred following stroke.In this population-based cohort study of patients admitted to hospital with stroke between 2008 and 2017 in Ontario, Canada, we used linked administrative data to calculate direct person-level costs in Canadian dollars in the one-year following stroke. We used a generalized linear model with a gamma distribution and a log link function to compare costs in women and men with and without adjustment for baseline clinical differences. We also assessed for an interaction between age and sex using restricted cubic splines to model the association of age with costs.We identified 101,252 patients . Unadjusted costs following stroke were higher in women compared to men . However, after adjustment, women had 3% lower costs compared to men . The lower cost in women compared to men was most prominent among people aged over 85 years (p for interaction = 0.03). Women incurred lower costs than men in outpatient care and rehabilitation, but higher costs in complex continuing care, long-term care, and home care.Patterns of resource utilization and direct medical costs were different between men and women after stroke. Our findings inform public payers of the drivers of costs following stroke and suggest the need for sex-based cost-effectiveness evaluation of stroke interventions with consideration of costs in all care settings.The online version contains supplementary material available at 10.1186/s12913-021-06669-w. Stroke is a leading cause of disability globally. In addition to affecting the lives of patients and their caregivers, stroke imposes a significant economic burden on the healthcare system , 2. CompThis information is important for several reasons. First, as the value of evaluating stroke care and outcomes stratified by sex is being increasingly recognized, , 7 inforWe described the direct healthcare cost in men and women as well as the healthcare settings in which these costs were incurred in the year after a hospitalization for stroke in Ontario, Canada with and without adjustment for comorbidities and costs incurred in the year prior to stroke.In this population-based retrospective cohort study, we identified all adults admitted to an acute care hospital with a most responsible diagnosis of ischemic stroke or intracerebral hemorrhage between April 1st 2008 and March 31st 2017 in Ontario, Canada’s most populous province with 14 million people , 10. We The primary outcome was the direct person-level cumulative cost, calculated from the perspective of the government payer in the first 365 days after the date of the index hospital admission. We estimated the costs of services borne by the Ontario Ministry of Health and Long-Term Care,  the publDirect costs to the Ontario public payer of physician billing for health services, prescription drugs, and outpatient diagnostic or laboratory services were calculated for each patient. Home care costs were estimated using the average cost per hour. Long-term care costs were calculated based on the government’s per diem payment rate. Given costs of encounters in the emergency department, hospital, and complex continuing care settings depend on the intensity of resource utilization, each encounter in these settings was assigned a resource intensity weight associated with its case-mix group, allowing for the calculation of the weighted cost for each visit based on the intensity of use of drugs, procedures, tests, and personnel. Resource intensity weights were calculated using standard methods from the Canadian Institute for Health Information . Our metWe used a generalized linear model with a gamma distribution and a log link function to assess the association between sex and one-year costs following stroke with and without adjustment for covariates . The intWe identified 101,252 patients, of whom 49% were women. Baseline characteristics are described in Table Table Figure The relative cost ratio is the exponentiated regression coefficient of the generalized linear model with a gamma distribution and a log link function. The coefficients for each covariate are shown in Supplemental Table p < 0.001), but the hazard of death in the two groups was similar after adjustment for all covariates   or coron,914  or Differences in baseline characteristics between men and women likely accounted for some of the cost differences as the magnitude and direction of sex difference in costs changed after adjustment for baseline characteristics. We expected that the direct healthcare costs from the public government payer’s perspective would be lower in the patients with shorter survival after stroke, compared to those who survive longer. We showed that costs were lower in women compared to men regardless of survival. Costs were 2% lower in women than men among patients alive at 1 year and 10% lower in women among those who died , that could result in differential reliance on publicly funded care. Future work on the reasons for sex differences in cost with information on quality indicators of care is needed.Our findings of differences in healthcare utilization patterns highlight the importance of considering all care settings when evaluating the cost-effectiveness of stroke interventions in men and women. For example, a treatment that reduces the need for long-term care may have more impact on reducing costs in women than men. Our work also informs the planning of bundled payment models to ensure that care needs are appropriately reimbursed and shifts in costs from one health setting to another are avoided , 27.In addition, we reported on different predictors of costs following stroke. Higher stroke severity, frailty, and intracerebral hemorrhage were associated with higher costs, which is consistent with prior work and our understanding of the disease , 28. We The current study has several strengths. Canada’s single-payer universal healthcare allowed us to examine individual-level direct costs in different healthcare settings that reflect the continuum of stroke care. We also considered pre-stroke costs which are not routinely included in health economic evaluations of stroke,  but thisThere are nevertheless several limitations worth discussing. First, we evaluated cost from the perspective of the Ontario public healthcare payer and could not account for costs incurred by individuals and their families, private insurers, nor societal costs such as income loss or other opportunity costs . Second,We quantified the sex differences in healthcare costs of stroke and found the settings where these costs were incurred were different between men and women. Future research on the mediators of cost differences in women and men after stroke, including how much of the costs are related to differences in post-stroke mortality and disability, is needed to understand the efficiency of healthcare expenditures and to optimise patient outcomes.Additional file 1.


Background: Nipah virus (NiV) is a zoonotic virus , which can also be transmitted through contaminated food or directly between people. According to a World Health Organization (WHO) report, the transmission of Nipah virus infection varies from animals to humans or humans to humans. The case fatality rate is estimated at 40% to 75%. The most infected regions include Cambodia, Ghana, Indonesia, Madagascar, the Philippines, and Thailand. The Nipah virus model is categorized into four parts: susceptible (S), exposed (E), infected (I), and recovered (R). Methods: The structural properties such as dynamical consistency, positivity, and boundedness are the considerable requirements of models in these fields. However, existing numerical methods like Euler–Maruyama and Stochastic Runge–Kutta fail to explain the main features of the biological problems. Results: The proposed stochastic non-standard finite difference (NSFD) employs standard and non-standard approaches in the numerical solution of the model, with positivity and boundedness as the characteristic determinants for efficiency and low-cost approximations. While the results from the existing standard stochastic methods converge conditionally or diverge in the long run, the solution by the stochastic NSFD method is stable and convergent over all time steps. Conclusions: The stochastic NSFD is an efficient, cost-effective method that accommodates all the desired feasible properties. In September 1998, in a village near Ipoh City, Perak State, West Malaysia, a case was reported as having similar symptoms to Japanese B encephalitis (JE) virus; it was dealt with as a standard routine case. Still, cases continued to occur in the region until February 1999. They were ascribed to the JE virus, which had previously caused porcine-associated outbreaks in Malaysia and had been treated accordingly. It was later discovered in March 1999 by virologists from the University of Malaya that the cases were caused by a new virus that belongs to the family Paramyxoviridae, which does not include the JE virus. The virus was named the “Nipah Virus (NiV)”, after Kampung Sungai Nipah (Nipah river village), whose patient’s specimens yielded the first viral isolates. Nipah virus is a zoonotic virus, which means it spreads between animals and people. It belongs to the Paramyxoviridae family and, genetically, it is related to the Hendra Virus.Flying foxes such as fruit bats are considered animal reservoirs for NiV. The outbreak of NiV infection started in September 1998 in a village near Ipoh City of Perak State, West Malaysia, which affected several areas just outside the town. In this epidemic, 27 patients were reported, which caused 15 deaths. Until March 1999, published sources quoted a prevalence of 265 cases of acute NiV encephalitis with 105 fatalities in Malaysia, giving a mortality of nearly 40%. In response to NiV, more than 1 million pigs were culled as it was discovered that sick pigs were significant carriers of the virus and further could transmit it to humans.Furthermore, it was discovered that some of the pigs might have consumed partially eaten fruit by fruit bats that were affected by the virus. The virus also hit Singapore, where sick animals were imported from Malaysia during the NiV outbreak. The total number of cases reported in Singapore was 11, out of which one death was documented. These two countries have had no issue reported since 1999, but outbreaks continue to occur in Bangladesh and India. The same kind of virus emerged in India and its neighboring country Bangladesh in 2001 but it was not investigated until March 2003. As Bangladesh is a Muslim country where the pig industry is not present, the virus was able to spread through raw date palm sap. Later, it was observed that affected bats’ saliva, urine, and excreta might have mixed in the fluid, which resulted in the spread of NiV. In 2014, the virus emerged in the Philippines, causing human deaths and sudden deaths in several horses. It was thought that the virus was transmitted to humans by way of direct exposure to infected horses.Tan et al. studied the incidence, distribution, and control of the Nipah virus . Chua pr1 represents the recovery rate of exposed individuals due to awareness; ε2 represents the recovery rate of infected individuals due to treatment; η represents the number of people quarantined; τ represents the number of isolation centers available; γ represents increased personal hygiene due to public awareness; σ represents a rate of public awareness; λ means surveillance coverage. The systematic flow of Nipah virus disease is presented in At any time, the states of the model are described as follows: S(t) represents people who are susceptible to the Nipah virus; E(t) means people exposed to the Nipah virus but not infected; I(t) means people who are infected with the Nipah virus and can transmit the virus; R(t) means people recovered from the Nipah virus; Λ represents the number of people susceptible based on the birth rate; β represents the recruitment rate; α represents progression rate of infected people; δ represents the death rate because of disease; µ represents a natural rate of death; εS (0) ≥ 0, E (0) ≥ 0, I (0) ≥ 0, R (0) ≥ 0, and The nonlinear ordinary differential equations by using the law of mass action are as follows:In this section, we will discuss the positivity and boundedness of solutions of the system (1)–(4) with initial conditions.Theorem 1.The results of the system (1)–(4) with given initial conditions are positive for allProof. By considering Equation (1),Theorem 2.The solutionsof the system (1)–(4) are bounded at any time andProof. →∞By considering the population function as follows:The system (1)–(4) admits two types of equilibria as follows: disease-free equilibrium = The next-generation matrix method is presented for the system (1)–(4). We calculate two types of matrices. One is a transition matrix, and the second is a transmission matrix, as follows:Theorem 3.The disease-free equilibrium =is locally asymptotically stable (LAS) when< 1.Proof. The Jacobian matrix at the disease-free equilibrium is as follows:By using the Routh–Hurwitz criteria of 2nd order Theorem 4.The endemic equilibrium =  is locally asymptotically stable (LAS) when> 1.Proof. The Jacobian matrix at the endemic equilibrium is as follows:By applying the Routh–Hurwitz Criterion for the 3rd order, Definition 1.Probability Space : A probability space is a three-tuple, , in which the three components are: Sample space: A nonempty setcalled the sample space, which represents all possible outcomes; Event space: A collectionof subsets ofcalled the event space. Ifis discrete, then usuallyIfis continuous, thenis usually a sigma-algebra on, and Probability function: A function,, that assigns probabilities to the events in. This will sometimes be referred to as a probability distribution over. The probability function,must satisfy several basic axioms:(i) (ii) (iii) Definition 2.Brownian Motion: The Brownian motion processis categorized by four facts .Ex.ExCt=StThe stochastic differential equations (SDEs) of pine wilt epidemic model (1)–(4) can be written asThe Euler–Maruyama method is used to determine the numerical result of Equation (6) by using the parameters’ values given in The Euler–Maruyama method of stochastic differential Equation (6) is as follows:The graphical behavior of the Euler–Maruyama scheme for both equilibria is presented as shown in In this section we introduce the non-parametric parameter into the system (1)–(4) as follows ,36:(8)dSConsider In addition, denote In addition,If “L” acts on a function Definition 3.Letbe a Brownian motion andbe an Ito drift-diffusion process that satisfies the stochastic differential equation:Ifthenis also an Ito drift-diffusion process, which satisfies as follows:Theorem 5.A unique solution (of the system (8)–(11) lies inwith initial conditions:Proof. By Ito’s formula, (8)–(11) admit positive solution in the sense of unique local on Next, we shall prove that the system (8)–(11) model admits Let A sequence at stopping times Since Then, Define a function Using Ito’s formula on (18), we have:For simplicity, we let The Set For every For “This section deals with well-known methods like the stochastic Runge–Kutta, and the proposed stochastic NSFD method with the given non-negative initial conditions as follows:The stochastic Runge–Kutta method could be developed on the system (12)–(15) as follows:Stage 1Stage 2Stage 3Stage 4Final stage:The stochastic NSFD could be developed for the system (8)–(11) as follows:Theorem 6.The stochastic NSFD method is stable if the eigenvalues of Equations (28)–(31) lie in the same unit circle for anyProof. Considering the functions from the system (28)–(31), we haveLemma 1.For the quadratic equation:,, 3, if and only if the following conditions are satisfied:(i) (ii) (iii) Proof. The proof is straightforward. □A comparison of the stochastic NSFD method with other stochastic numerical methods is presented. It is easy to see that other stochastic numerical methods conditionally converge or diverge with larger time step values by looking at the numerical solutions, as shown in h = 100. Furthermore, Through this study, we investigated the transmission dynamics of the Nipah virus in humans. The whole manuscript comprises three Sections. Modeling, terminology related to epidemiology, and Nipah virus are the critical points of The stochastic non-standard finite difference scheme is designed for the communication dynamics of the Nipah virus. Unfortunately, the methods mentioned earlier, like Euler–Maruyama and stochastic Runge–Kutta of order 4th, are unsuitable because they depend on time step size. Thus, Euler–Maruyama and stochastic Runge–Kutta are tentatively convergent. When we increase the time step size, the graph of Euler–Maruyama and stochastic Runge–Kutta gives variation in results from time to time that they display as divergent. Furthermore, the existing numerical methods did not preserve the structure of the continuous model. Thus, these are the gaps in the literature that need to be filled. For this reason, we have introduced the non-standard finite difference method, which preserves the actual structure of the continuous model, such as positivity, boundedness, and dynamical consistency. The new well-known numerical scheme—such as the stochastic non-standard finite difference scheme—is independent of time step size. The SNSFD scheme is a comfortable tool on behalf of dynamical properties like stability, positivity, and boundedness and shows the exact behavior of the continuous model. In the future, we will extend the idea used in this work to different types of modelling, including spatiotemporal, fractional, fractal fractional, and delay problems of dynamical systems.