Dual effects of ipecac alkaloids with potent antiviral activity against foot-and-mouth disease virus as replicase inhibitors and direct virucides.

Foot-and-Mouth Disease (FMD) is a contagious, blistering disease caused by the Foot-and-Mouth Disease virus (FMDV), which affects livestock globally. Currently, no commercial antiviral agent is available for effective disease control. This study investigated the antiviral potential of natural-derived alkaloids against FMDV in BHK-21 cells. Twelve alkaloids were assessed for their antiviral activities at various stages of FMDV infection, including pre-viral entry, post-viral entry, and prophylactic assays, as well as attachment and penetration assays by evaluating cytopathic effect reduction and directed-virucidal effects. The results showed that ipecac alkaloids, cephaeline (CPL) and emetine (EMT), exhibited dual effects with robust antiviral efficacy by reducing cytopathic effect and inhibiting FMDV replication in a dose-dependent manner. Evaluation through immunoperoxidase monolayer assay and RT-PCR indicated effectiveness at post-viral entry stage, with sub-micromolar EC50 values for CPL and EMT at 0.05 and 0.24 µM, respectively, and high selective indices. Prophylactic effects prevented infection with EC50 values of 0.23 and 0.64 µM, respectively. Directed-virucidal effects demonstrated significant reduction of extracellular FMDV, with CPL exhibiting a dose-dependent effect. Furthermore, the replicase (3Dpol) inhibition activity was identified using the FMDV minigenome assay, which revealed strong inhibition with IC50 values of 0.15 µM for CPL and 4.20 µM for EMT, consistent with the decreased negative-stranded RNA production. Molecular docking confirmed the interaction of CPL and EMT with residues in the active site of FMDV 3Dpol. In conclusion, CPL and EMT exhibited promising efficacy through their dual effects and provide an alternative approach for controlling FMD in livestock.

Enhanced production of recombinant coxsackievirus A16 using a serum-free HEK293A suspension culture system for bivalent enterovirus vaccine development.

Coxsackievirus A16 (CVA16) is one of the primary pathogens that causes hand, foot, and mouth disease (HFMD) in young children. In previous studies, CVA16 vaccine development has encountered several challenges, such as inefficient replication of the CVA16 virus in present culture systems, the induction of only mild neutralizing antibody titers, and neutralizing antibodies induced by certain vaccine candidates that are unable to protect against CVA16 viral challenge. In this study, we constructed a DNA-launched CVA16 infectious clone (CVA16ic) based on the genomic sequence of the CVA16 N5079 strain to minimize interference from viral quasispecies. The biochemical properties of this CVA16ic strain were similar to those of its parental strain. Serum-free HEK293A suspension cells, which produced higher virus titers than Vero cells, were demonstrated to improve CVA16 production yields. In addition, our study showed that inactivated EV-A71 antigens could enhance the immunogenicity of inactivated CVA16 mature/full particles (F-particles), suggesting that a bivalent CVA16 and EV-A71 vaccine may be an effective strategy for CVA16 vaccine development. These findings are expected to provide novel strategies and accelerate the development of bivalent HFMD vaccines.

The isolation and serotyping of foot-and-mouth disease virus in Iran during 2019-2022.

Foot-and-mouth disease (FMD) is a significant transboundary animal disease that has a considerable economic impact on livestock systems worldwide. In order to determine the presence and type of FMD virus in Iran, a total of 90 samples of vesicular fluid and epithelial tissue were collected from the tongues, tooth pads, and hooves of clinically suspect cattle on 40 vaccinated farms in 9 provinces of Iran. These samples were collected during four years, from January 2019 to December 2022, and the vaccine was a locally produced polyvalent inactivated vaccine. The collected samples were analyzed using ELISA and isolation methods to identify and characterize the FMD virus. The results of the ELISA tests revealed that 66.66% of the samples were positive for FMD, and the serotypes of the virus were determined. Considering ELISA reslut, 62% of the samples were assigned to serotype O, 33% to serotype A, and 5% to serotype Asia-1. Furthermore, 90% of the positive samples were inoculated onto monolayer cultures of pig kidneys (IB-RS2) for isolation and antigen detection by serotype-specific ELISA kit. The great majority of detected serotype O viruses were from Esfahan province, while the most detected serotype A and serotype Asia-1 viruses were from Qom and Tehran provinces, respectively. These findings indicate that the ELISA and isolation methods are suitable for identifying and typing FMD viruses. The vaccination program in Iran, which includes three serotypes (O, A, and Asia-1), appears to be effective in controlling the spread of the disease. However, the continued circulation of these serotypes in most provinces suggests that ongoing surveillance and vaccination efforts are necessary.

Analysis of the immunological response elicited by a polyvalent foot and mouth disease vaccine and its compatibility with a diva test in Jimma Town, Ethiopia.

The research was conducted in Jimma town, Oromiya Regional State, from October 2022 to June 2023, with the aim of assessing the immune response of polyvalent FMD (Foot and Mouth Disease) vaccine. The study involved 34 cattle in a longitudinal study, divided into two groups: 29 vaccinated and 5 unvaccinated. The vaccinated cattle received an inactivated polyvalent FMD virus vaccine produced by the National Veterinary Institute. Blood samples were collected on days 0, 14, 21, 35, 80, and 125 after vaccination and tested using Virus Neutralization Test and 3ABC ELISA. The results showed a significant increase in neutralizing antibodies against structural proteins in all vaccinated cattle on day 14 after vaccination for all three serotypes. (A/ETH/21/2000, p = 0.015; O/ETH/38/2005, p = 0.017; SAT2/ETH/64/2009, p = 0.007). On day, fourteen of post-vaccination vaccinated group showed immune response equal or above 1.5 log10 in a proportion of 69%, 73% and 94% for serotype A/ETH/21/2000, O/ETH/38/2005 and SAT2/ETH/64/2009 respectively. The status of raised antibody titer on day 125 post-vaccination showed decreasing by 14%, 18% and 4% for serotype A/ETH/21/2000, O/ETH/38/2005 and SAT2/ETH/64/2009 respectively. The DIVA test, or 3ABC ELISA, used to differentiate infected from vaccinated animals, revealed the absence of immune response to the Non-structural protein in the vaccinated cattle group. Conversely, the unvaccinated group showed no recorded antibody titer to both structural and non-structural proteins. In summary, the commercially available FMD vaccine, comprising serotype A, O, and SAT2, triggers an immune response to the structural protein rather than the non-structural protein after the initial administration. This outcome implies that FMD vaccines from the National Veterinary Institute align with the DIVA test. Nevertheless, additional efforts may be necessary to bolster the strength and duration of the vaccine-induced immune response.

Evaluation and comparison of performances of six commercial NSP ELISA assays for foot and mouth disease virus in Thailand.

ELISA kits that detect antibodies to the non-structural protein (NSP) of the foot-and-mouth disease virus (FMDV), commonly referred to as NSP-ELISA, can distinguish between vaccinated and naturally infected animals. They can play an essential role in demonstrating 'proof-of-freedom' during the control of FMD. Although various NSP-ELISA kits are available in Thailand, information regarding their performance is lacking. To select the most appropriate NSP-ELISA kit for our specific purpose, we must compare their performance using carefully characterized sera. This will ensure that we maximize the benefits of our testing. In this study, six NSP-ELISA kits sold in Thailand-Biovet, ID Screen, VDPro, IDEXX, PrioCHECK, and KUcheck-F-were evaluated and compared. A total of 800 serum samples were examined, including samples from 357 cattle and 29 buffaloes in outbreak areas, as well as 14 swine serum samples from the Vaccine Quality Control Unit of the Bureau of Veterinary Biologics, Ministry of Agriculture and Cooperation, Thailand. Four hundred samples were confirmed to originate from animals infected with FMDV through ELISA typing (n = 11, tested as representative samples in each farm) and/or RT-PCR (n = 400, all samples), serving as positive control sera. Additionally, 400 negative control sera were obtained from Japan (97 cattle and 300 pigs) and Australia (3 goats), certified by the World Organisation for Animal Health as 'free of FMD'. The sensitivity and specificity of the six tests were determined based on the results obtained from two-by-two tables. Cohen's kappa statistics were calculated for the six tests to assess their concordance, and the diagnostic accuracy of the assays was also determined. For all six NSP-ELISA kits, the sensitivity ranged from 97.75 to 99.50%, and the specificity ranged from 97.25 to 100%. Cohen's kappa statistics ranged from 0.96 to 1.00, and diagnostic accuracy ranged from 98.13 to 99.75%. The study results indicated that the test kits have statistically similar sensitivity, specificity, concordance, and diagnostic accuracy, suggesting they can be used interchangeably. However, ID Screen demonstrated the highest sensitivity and specificity among all kits tested. Therefore, if a single kit were to be selected from the six evaluated, ID Screen would be the most appropriate choice. These findings can aid in selecting the most suitable test kit. Therefore, it is recommended to consider purchasing a diverse range of effective test kits. Furthermore, these findings can provide guidance for expanding the use of test kits, particularly with the growing availability of NSP-ELISA kits in the market.

An evaluation of the efficiency and effectiveness of diagnostic tests for foot and mouth disease: are novel diagnostic tests for FMD more feasible than conventional tests in Southeast Asia?

Foot and mouth disease (FMD) remains endemic in many areas of continental Southeast Asia (SEA). It is responsible for substantial economic losses in the smallholder sector and threatens livelihoods. In recent years, novel diagnostic tests have been developed which reportedly detect FMD virus more effectively and efficiently. This critically appraised topic (CAT) aimed to evaluate the feasibility of these diagnostic tests for FMD in SEA compared to conventional tests. Relevant studies that evaluate diagnostic tests are identified and critically assessed, and recommendations are made on suitable potential diagnostic tests for use in the smallholder sector in SEA. A systematic search of electronic databases (CABI: CAB Abstracts, Web of Science Core Collections) was carried out to identify relevant studies that compared novel and conventional diagnostic tests. The search strategy initially identified 12 papers, of which six fulfilled all the inclusion criteria and were selected for this review. Most of the selected studies had limitations in design and comparability, making it difficult to validly compare the effectiveness and efficiency of the relevant diagnostic tests. These limitations include variation in sample characteristics, methodology, measurable outcomes and the different aspects of the diagnostic tests that each study focused on. Most studies concluded that novel diagnostic tests were more effective and efficient than conventional tests: had greater analytical sensitivity and specificity, were more robust, had a wider range of processable sample types and serotypes, could detect various diseases, had faster testing speeds and provided greater value for money. However, strong recommendations on which specific diagnostic test to rely on could not be made, since there was conflicting evidence and multiple confounding factors. Overall, the evidence found did not entirely apply to the target scenario, being SEA smallholder farms. Recommendations for the target scenario were also made based on the study findings.

Epidemiological characterization of hand, foot, and mouth disease among hospitalized children from 2014 to 2023 in a hospital in Henan Province: Longitudinal surveillance study.

Hand, foot, and mouth disease (HFMD) is an acute infectious illness primarily caused by enteroviruses. The present study aimed to describe the epidemiological characteristics of hospitalized HFMD patients in a hospital in Henan Province (Zhengzhou, China), and to predict the future epidemiological parameters. In this study, we conducted a retrospective analysis of general demographic and clinical data on hospitalized children who were diagnosed with HFMD from 2014 to 2023. We used wavelet analysis to determine the periodicity of the disease. We also conducted an analysis of the impact of the COVID-19 epidemic on the detection ratio of severe illness. Additionally, we employed a Seasonal Difference Autoregressive Moving Average (SARIMA) model to forecast characteristics of future newly hospitalized HFMD children. A total of 19 487 HFMD cases were included in the dataset. Among these cases, 1515 (7.8%) were classified as severe. The peak incidence of HFMD typically fell between May and July, exhibiting pronounced seasonality. The emergence of COVID-19 pandemic changed the ratio of severe illness. In addition, the best-fitted seasonal ARIMA model was identified as (2,0,2)(1,0,1)12. The incidence of severe cases decreased significantly following the introduction of the vaccine to the market (χ2 = 109.9, p < 0.05). The number of hospitalized HFMD cases in Henan Province exhibited a seasonal and declining trend from 2014 to 2023. Non-pharmacological interventions implemented during the COVID-19 pandemic have led to a reduction in the incidence of severe illness.

The Relevance of Bacillary Layer Detachment and Choroidal flow Insufficiency in Unilateral Acute Idiopathic Maculopathy Associated with Hand, Foot, and Mouth Disease: A Case Report.

In a case of unilateral acute idiopathic maculopathy (UAIM) following hand, foot, and mouth disease, we aim to discuss the decreased perfusion of choriocapillaris secondary to systemic inflammation as shown by optical coherence tomography angiography (OCTA) and to assess the prognostic significance of bacillary layer detachment (BALAD). A 33-year-old male presented with a decrease of vision in the right eye (OD) for 5 days preceding viral prodromal symptoms and vesicular lesions on bilateral palms and soles along with vesicles and ulcers on the oral mucosa. The best-corrected visual acuity was finger counting at 1 meter distance in OD and 20/20 in his left eye (OS). Dilated fundus examination revealed a circular white-grey dome-shaped elevated lesion at the macula indicative of serous retinal detachment in OD. Spectral-domain optical coherence tomography demonstrated BALAD associated with adjacent subretinal and intraretinal fluid along with pigment epithelium detachment and disruption of ellipsoid and interdigitation zones. OCTA showed decreased choriocapillaris perfusion. All the investigations were normal in OS. The resolution of BALAD occurred during the first 2 days, which was followed by gradual improvement of choriocapillaris flow that lasted 2 months. UAIM is associated with hand, foot, and mouth disease. OCTA demonstrates both qualitative and quantitative data by detecting alterations in the choriocapillaris flow, which could be monitored during the disease course.

Analysis of miRNAs involved in mouse brain injury upon Coxsackievirus A6 infection.

Introduction: Coxsackievirus A6 (CV-A6) has emerged as the predominant epidemic strain responsible for hand, foot and mouth disease (HFMD). CV-A6 infection can result in severe clinical manifestations, including encephalitis, meningitis, and potentially life-threatening central nervous system disorders. Our previous research findings demonstrated that neonatal mice infected with CV-A6 exhibited limb weakness, paralysis, and ultimately succumbed to death. However, the underlying mechanism of CV-A6-induced nervous system injury remains elusive. Numerous reports have highlighted the pivotal role of miRNAs in various viral infections. Methods: Separately established infection and control groups of mice were used to create miRNA profiles of the brain tissues before and after CV-A6 transfection, followed by experimental verification, prediction, and analysis of the results. Results: At 2 days post-infection (dpi), 4 dpi, and 2dpi vs 4dpi, we identified 175, 198 and 78 significantly differentially expressed miRNAs respectively using qRT-PCR for validation purposes. Subsequently, we predicted target genes of these differentially expressed miRNAs and determined their potential targets through GO (Gene Ontology) enrichment analysis and KEGG (Kyoto Encyclopedia of Genes and Genomes) enrichment analysis. Finally, we verified the miRNA-mRNA pairing via double luciferase experiments while confirming functional enrichment of target genes through Western Blotting analyses. Discussion: The results from this study suggest that transcriptional regulation, neuronal necrosis, pro-inflammatory cytokine release, and antiviral immunity are all implicated in the pathogenesis of central nervous system injury in mice infected with CV-A6. Brain injury resulting from CV-A6 infection may involve multiple pathways, including glial cell activation, neuronal necrosis, synaptic destruction, degenerative diseases of the nervous system. It can even encompass destruction of the blood-brain barrier, leading to central nervous system injury. The dysregulated miRNAs and signaling pathways discovered in this study provide valuable insights for further investigations into the pathogenesis of CV-A6.

The key mechanisms of multi-system responses triggered by central nervous system damage in hand, foot, and mouth disease severity.

Hand, foot, and mouth disease (HFMD) is a prevalent infectious affliction primarily affecting children, with a small portion of cases progressing to neurological complications. Notably, in a subset of severe HFMD cases, neurological manifestations may result in significant sequelae and pose a risk of mortality. We systematically conducted literature retrieval from the databases PubMed (1957-2023), Embase (1957-2023), and Web of Science (1957-2023), in addition to consulting authoritative guidelines. Subsequently, we rigorously selected the most relevant articles within the scope of this review for comprehensive analysis. It is widely recognized that the severity of HFMD is attributed to a multifaceted array of pathophysiological mechanisms. The implication of multi-system dysfunction appears to be perturbances of the human defense system; therefore, it contributes to the severity of HFMD. In this review, we provide an overview and analysis of recent insights into the molecular mechanisms contributing to the severity of HFMD, with a particular focus on cytokine release syndrome, the involvement of the renin-angiotensin system, regional immunity, endothelial dysfunction, catecholamine storm, viral invasion, and the molecular mechanisms of neurological damage. We speculate that the domino effect of diverse physiological systems, initiated by damage to the central nervous system, serve as the primary mechanisms governing the severity of HFMD. Simultaneously, we emphasize the knowledge gaps and research urgently required to delineate a quick roadmap for ongoing and essential studies on HFMD.

Effect of GnRH treatment as a potential solution for ovarian disorders in dairy cows infected with foot and mouth disease in Indonesian smallholder farms.

Background: The outbreak of foot and mouth disease (FMD) in Indonesia induces reproductive disorders in dairy cows that lead to economic losses to smallholder dairy farms. Aim: The study was to assess the influence of FMD on reproductive traits and evaluate the effect of gonadotropin hormone-releasing hormone (GnRH) administrations on the reproductive performance in FMD-infected dairy cows. Methods: The study was conducted in Jemowo village, Taman Sari sub-district, Boyolali district, Central Java, Indonesia. A total of 155 cows were used to identify the reproductive disorders on FMD-infected dairy cows aged 2-10 years old. Cows were raised in similar conditions and fed diets. A single dose of 2 ml GnRH was injected intramuscularly into 96 ovarian disorder cows. Reproductive performance was measured by service per conception (S/C), conception rate (CR), and pregnancy rate (PR). A descriptive study was conducted to demonstrate the results. Results: The study showed that 61.9% of FMD-infected cows had reproductive disorders, whereby 53.5% ovarian hypofunction, 4.52% silent heat, 1.94% repeat breeder, 1.29% ovarian atrophy, and 0.65% endometritis. FMD-infected cows injected with GnRH had a 98% reproductive recovery rate. Moreover, the S/C, CR, and PR of cows injected with GnRH were 2.02%, 51%, and 85%. Conclusion: GnRH administrations enhanced the reproductive traits of FMD-infected dairy cows indicated by the improvement of CR and PR.

Phylogenetic analyses and antigenic characterization of foot-and-mouth disease virus PanAsia lineage circulating in China between 1999 and 2023.

Foot-and-mouth disease (FMD) is one of the most important transboundary animal diseases caused by foot-and-mouth disease virus (FMDV), leading to significant economic losses worldwide. The first report of PanAsia lineage of FMDV in China was in 1999. Since 2011, 18 outbreaks attributed to PanAsia lineage viruses have been reported across 7 provinces or municipality in China. Phylogenetic analysis indicated that these PanAsia strains were clustered into three distinct clades (clade 1, clade 2, and clade 3), with nucleotide homology ranging from 91.4% to 100%. The outbreaks of FMD caused by clade 1 strains occurred around 1999 when this lineage was prevalent globally. Clade 2 strains dominated from 2011 to 2013, while clade 3 strains were prevalent during 2018-2019, sharing only 93% homology with clade 2 strains and 91% with clade 1 strains. Tracing analysis showed that these outbreaks represented 3 distinct introductions of PanAsia viruses into China. Virus neutralization tests (VNT) have demonstrated that current commercial vaccines are effective to protect susceptible animals against these strains (r1 > 0.3). However, the growing demand for livestock has promoted animal movement and encouraged the exchange of products, services, and materials between countries, thereby heightening the risk of exotic strain incursions. Therefore, it is imperative to reinforce border controls and limit animal movements among various Asian countries continually to reduce the risk of new transboundary diseases, such as FMD incursion. Additionally, PanAsia-2 strains need to be taken seriously to prevent its incursions, and the relevant vaccines against PanAsia-2 strains needs to be stockpiled in preparation for any possible incursion.

Evolving pathogen trends and spatial-temporal dynamics of hand, foot, and mouth disease in Fengxian District, Shanghai (2009-2022).

Hand, foot, and mouth disease (HFMD) is a prevalent acute infectious disease caused by enteroviruses, presenting substantial public health challenges in Shanghai, especially among children. The dynamic nature of HFMD's etiology necessitates an ongoing evaluation of its epidemiological and virological trends to inform effective control strategies. This study aims to investigate the epidemiological patterns and viral evolution of HFMD in Fengxian District, Shanghai, China, with a focus on shifts in predominant viral strains over a 14-year period. We conducted a retrospective analysis of HFMD cases reported to the National Notifiable Disease Reporting System in Fengxian District from January 1, 2009 to December 31, 2022. Epidemiological trends, strain prevalence, and demographic impacts were assessed. A total of 27,272 HFMD cases were documented during the study period, with incidence showing pronounced seasonal fluctuations-peaking in spring and summer and a lesser peak in autumn. The disease incidence demonstrated significant positive correlations with several meteorological variables: daily average temperature (r = 0.30, P < 0.05), relative humidity (r = 0.20, P < 0.05), wind speed (r = 0.17, P < 0.05), and precipitation (r = 0.17, P < 0.05). Geographically, Nanqiao Town, Fengcheng Town, and Xidu Subdistrict reported the highest incidence rates. The demographic analysis revealed a male-to-female ratio of 1.60:1, predominantly affecting children aged 1-3 years. Prior to 2017, Enterovirus 71 (EV71) and Coxsackievirus A16 (CoxA16) were the primary detected strains; post-2017, Coxsackievirus A6 (CoxA6) emerged as the dominant strain. Statistical analysis confirmed significant year-to-year variations in virus detection rates, with decreasing trends for EV71 and other enteroviruses and an increasing trend for CoxA6. The findings indicate a distinct seasonal incidence of HFMD in Fengxian District. This study underscores the need for targeted public health education, enhanced surveillance, and proactive measures in childcare facilities to mitigate disease spread during peak seasons. Moreover, the evolving viral landscape warrants accelerated efforts in vaccine development against new strains to reduce HFMD incidence.

Exploring the Birthday Week Effect on Hand, Foot, and Mouth Disease in Yunnan Province, China, From 2008 to 2022: Surveillance Data Analysis.

Background: Hand, foot, and mouth disease (HFMD) is a notable infectious disease predominantly affecting infants and children worldwide. Previous studies on HFMD have primarily focused on natural patterns, such as seasonality, but research on the influence of important social time points is lacking. Several studies have indicated correlations between birthdays and certain disease outcomes. Objective: This study aimed to explore the association between birthdays and HFMD. Methods: Surveillance data on HFMD from 2008 to 2022 in Yunnan Province, China, were collected. We defined the period from 6 days before the birthday to the exact birthday as the "birthday week." The effect of the birthday week was measured by the proportion of cases occurring during this period, termed the "birthday week proportion." We conducted subgroup analyses to present the birthday week proportions across sexes, age groups, months of birth, and reporting years. Additionally, we used a modified Poisson regression model to identify conditional subgroups more likely to contract HFMD during the birthday week. Results: Among the 973,410 cases in total, 116,976 (12.02%) occurred during the birthday week, which is 6.27 times the average weekly proportion (7/365, 1.92%). While the birthday week proportions were similar between male and female individuals (68,849/564,725, 12.19% vs 48,127/408,685, 11.78%; χ21=153.25, P<.001), significant differences were observed among different age groups (χ23=47,145, P<.001) and months of birth (χ211=16,942, P<.001). Compared to other age groups, infants aged 0-1 year had the highest birthday week proportion (30,539/90,709, 33.67%), which is 17.57 times the average weekly proportion. Compared to other months, patients born from April to July and from October to December, the peak months of the HFMD epidemic, had higher birthday week proportions. Additionally, a decreasing trend in birthday week proportions from 2008 to 2022 was observed, dropping from 33.74% (3914/11,600) to 2.77% (2254/81,372; Cochran-Armitage trend test: Z=-102.53, P<.001). The results of the modified Poisson regression model further supported the subgroup analyses findings. Compared with children aged >7 years, infants aged 0-1 year were more likely to contract HFMD during the birthday week (relative risk 1.182, 95% CI 1.177-1.185; P<.001). Those born during peak epidemic months exhibited a higher propensity for contracting HFMD during their birthday week. Compared with January, the highest relative risk was observed in May (1.087, 95% CI 1.084-1.090; P<.001). Conclusions: This study identified a novel "birthday week effect" of HFMD, particularly notable for infants approaching their first birthday and those born during peak epidemic months. Improvements in surveillance quality may explain the declining trend of the birthday week effect over the years. Higher exposure risk during the birthday period and potential biological mechanisms might also account for this phenomenon. Raising public awareness of the heightened risk during the birthday week could benefit HFMD prevention and control.

Stearoyl coenzyme A desaturase 1 (SCD1) regulates foot-and-mouth disease virus replication by modulating host cell lipid metabolism and viral protein 2C-mediated replication complex formation.

The life cycle of foot-and-mouth disease virus (FMDV) is tightly regulated by host cell lipid metabolism. In previous studies, we reported downregulated expression of stearoyl coenzyme A desaturase-1 (SCD1), a key enzyme of fatty acid metabolism, in BHK-VEC cells (a virus-negative cell line derived from BKH-21 cells with persistent FMDV infection) on comparing transcriptomic data for BHK-VEC and BHK-21 cells (Y. Yuan et al., Front Cell Infect Microbiol 12:940906, 2022, https://doi.org/10.3389/fcimb.2022.940906; L. Han et al., Vet Microbiol 263:109247, 2021, https://doi.org/10.1016/j.vetmic.2021.109247). In the present study, we identify that SCD1 regulates FMDV replication. SCD1 overexpression or exogenous addition of oleic acid (OA), a product of the enzymatic activity of SCD1, increased FMDV replication in both BHK-21 cells and SCD1-knockdown cells. Overexpression of SCD1 or exogenous addition of OA restored FMDV infection and replication in BHK-VEC cells, and OA also promoted FMDV replication in BHK-21 cells with persistent FMDV infection. SCD1 recruited the nonstructural FMDV protein 2C to a detergent-resistant membrane located in the perinuclear region of cells to form replication complexes. Inhibiting SCD1 enzyme activity resulted in a significantly decreased number of FMDV replication complexes with abnormal morphology. Inhibition of SCD1 activity also effectively decreased the replication of other RNA viruses such as respiratory enteric orphan virus-3-176, poliovirus-1, enterovirus 71, and vesicular stomatitis virus. Our results demonstrate that SCD1, as a key host regulator of RNA virus replication, is a potential target for developing novel drugs against infections by RNA viruses. IMPORTANCE: Many positive-stranded RNA viruses, including foot-and-mouth disease virus (FMDV), alter host membranes and lipid metabolism to create a suitable microenvironment for their survival and replication within host cells. In FMDV-infected cells, the endoplasmic reticulum membrane is remodeled, forming vesicular structures that rely heavily on increased free fatty acids, thereby linking lipid metabolism to the FMDV replication complex. Nonstructural FMDV protein 2C is crucial for this complex, while host cell enzyme stearoyl coenzyme A desaturase 1 (SCD1) is vital for lipid metabolism. We found that FMDV infection alters SCD1 expression in host cells. Inhibiting SCD1 expression or its enzymatic activity markedly decreases FMDV replication, while supplementing oleic acid, a catalytic product of SCD1, regulates FMDV replication. Additionally, SCD1 forms part of the FMDV replication complex and helps recruit 2C to a detergent-resistant membrane. Our study provides insights into the pathogenesis of FMDV and a potential novel drug target against the virus.

Protein characterization of an Indonesian isolate of foot and mouth disease virus inactivated with formaldehyde and binary ethylenimine.

Background and Aim: Foot-and-mouth disease (FMD) is a highly contagious viral disease of cloven-footed animals. It is a major threat to livestock production worldwide, causing significant economic losses. Inactivation of FMD virus (FMDV) is crucial for vaccine development and control of outbreaks. However, traditional inactivation methods can sometimes damage the viral protein, affecting vaccine efficacy. Therefore, finding new inactivating agents that effectively inactivate the virus while preserving the integrity of its proteins is an important research area. This study investigated the optimal materials (0.04% formaldehyde, 0.001 M binary ethylenimine [BEI], or a combination) for inactivating and preserving the specific molecular weight of Serotype O FMDV protein. Materials and Methods: This study used serotype O FMDV isolated from several areas of East Java. The virus was inoculated into baby hamster kidney-21 cells, and the titer was calculated using the TCID50 Assay. The virus was inactivated using 0.04% formaldehyde, 0.001 M BEI, or a combination of 0.04% formaldehyde and 0.001 M BEI. Inactive viral proteins were characterized using sodium dodecyl sulfate-polyacrylamide gel electrophoresis and western blotting. Results: Serotype O FMDV can be inactivated using 0.04% formaldehyde while preserving specific FMDV proteins, specifically VP0 and VP3 with a molecular weight (MW) of 36 kDa and VP3 with a MW of 24 kDa. Serotype O FMDV can be inactivated by 0.001 M BEI while preserving specific FMDV proteins, specifically VP0 with a MW of 35 kDa, VP3 with a MW of 28 kDa, and VP1 with a MW of 23 kDa. FMDV serotype O can be inactivated using a combination of 0.04% formaldehyde and 0.001 M BEI while preserving specific FMDV proteins, specifically VP0 and VP3 with a MW of 36 kDa and VP3 with a MW of 24 kDa. Conclusion: This study found that 0.04% formaldehyde, alone or in combination with 0.001 M BEI, was effective for inactivating and preserving the specific molecular weight of Serotype O FMDV protein. The limitation of this study was the inactivations of the virus have not yet been tested for their potency on experimental animals. Further research is warranted to investigate the inactivation kinetics of these materials, including their potency on experimental animals. Additionally, a comparison of the inactivation rates between 0.04% formaldehyde alone and the combination with BEI would help to determine the optimal inactivation agent for future applications.

Dendritic cell-based biomimetic nanoparticles for foot-and-mouth disease induce robust cellular immunity.

Foot-and-mouth disease (FMD) is a highly contagious and economically devastating viral disease of ruminants and swine, badly affecting the livestock industry worldwide. In clinical practice, vaccination is a frequently employed strategy to prevent foot-and-mouth disease (FMDV). However, commercial inactivated vaccines for FMD mainly rely on humoral immunity, exhibiting poor cellular immune responses and causing adverse reactions. Here, we use the double emulsion method to prepare poly (lactic-co-glycolic acid) nanoparticles (PLGA-NP) encapsulated with IL-2 cytokines, wrap the dendritic cell (DC) membrane carrying FMDV antigen information on the surface of the nanoparticles, obtaining a biomimetic nanoparticle vaccine Biom@DC with uniform size. This vaccine can effortlessly move through lymph nodes due to its nanoscale size advantage. It also possesses DC ability to present antigens, and antigen presentation can be made more effective with high biocompatibility. The sustained release of IL-2 encapsulated in the core of PLGA-NP in vivo can effectively promote the body's cellular immune response. Immune tests on mice have shown that Biom@DC may greatly increase T cell activation and proliferation both in vivo and in vitro, while also significantly reducing the fraction of inhibitory Treg cells. Furthermore, in the micro serum neutralization assay for FMDV, it has been demonstrated that the group vaccinated with Biom@DC exhibits a clear neutralizing effect. Given its strong immunogenicity, Biom@DC has the potential to develop into a novel, potent anti-FMDV vaccination.

Enhanced Effects of ISA 207 Adjuvant via Intradermal Route in Foot-and-Mouth Disease Vaccine for Pigs.

In South Korea, a mandatory nation-wide foot-and-mouth disease (FMD) vaccination policy is in place. However, a major side effect of the current method of intramuscular (IM) administration of oil-adjuvanted FMD vaccines is the formation of granulomas in the muscles of pigs. To address this issue, we assessed the possible application of intradermal (ID) vaccination. Initially, we compared the serological immune response in specific pathogen-free pigs inoculated with FMD vaccines formulated with eight different adjuvants, administered twice at the neck site using a syringe with a needle via the ID route. Among the formulations (water-in-oil-in-water (W/O/W), oil-in-water (O/W), and polymer nanomaterials), ISA 207 of W/O/W was the most effective in inducing immunogenicity followed by ISA 201 of W/O/W. ISA 207 was further tested in formulations of different antigen doses (12 or 1.2 µg) delivered via both IM and ID routes. All four treatments successfully protected the pigs against FMD virus challenges. To assess the feasibility of the field application of the vaccines with ISA 207, we conducted ID vaccination of conventional pigs using a needle-free device, resulting in the detection of significant levels of neutralizing antibodies. ISA 207 was shown to be superior to ISA 201 in inducing immunogenicity via the ID route. In conclusion, ISA 207 could be a suitable adjuvant for ID vaccination in terms of vaccine efficacy for FMD, allowing for alternate use of ID vaccination and subsequent reduction in the incidences of granuloma formation in the field.

Long-Term Immunogenicity Study of an Aluminum Phosphate-Adjuvanted Inactivated Enterovirus A71 Vaccine in Children: An Extension to a Phase 2 Study.

Enterovirus A71 (EV-A71) causes hand, foot, and mouth disease in infants and children with potential for fatal complications such as encephalitis and acute flaccid myelitis. This study examined the long-term immunity conferred by EV71vac, an inactivated EV-A71 vaccine adjuvanted with aluminum phosphate, in children from the age of 2 months to <6 years, for up to 5 years after the first immunization. A total of 227 participants between 2 months and <6 years of age who had previously received either EV71vac or placebo in the phase two clinical study were enrolled. Subjects were divided into age groups: 2 years to <6 years (Group 2b), 6 months to <2 years (Group 2c), and 2 months to <6 months (Group 2d). At Year 5, the neutralizing antibody titers against the B4 subgenotype remained high at 621.38 to 978.20, 841.40 to 1159.93, and 477.71 to 745.07 for Groups 2b, 2c, and 2d, respectively. Cross-neutralizing titers at Year 5 remained high against B5 and C4a subgenotypes, respectively. No long-term safety issues were reported. Our study provides novel insights into the long-term immunity conferred by EV71vac in children aged from two months to six years, particularly in those who received EV71vac between two and six months of age.

Virulence and Immune Evasion Strategies of FMDV: Implications for Vaccine Design.

Foot-and-mouth disease (FMD) is globally recognized as a highly economically devastating and prioritized viral disease affecting livestock. Vaccination remains a crucial preventive measure against FMD. The improvement of current vaccine platforms could help control outbreaks, leading to the potential eradication of the disease. In this review, we describe the variances in virulence and immune responses among FMD-susceptible host species, specifically bovines and pigs, highlighting the details of host-pathogen interactions and their impact on the severity of the disease. This knowledge serves as an important foundation for translating our insights into the rational design of vaccines and countermeasure strategies, including the use of interferon as a biotherapeutic agent. Ultimately, in this review, we aim to bridge the gap between our understanding of FMDV biology and the practical approaches to control and potentially eradicate FMD.