Application of the Japanese Verbal Learning Test to Patients With Alzheimer's Disease in the Early Stage.

INTRODUCTION: This study aimed to investigate the mechanism of memory function in the context of explicit memory in early-stage Alzheimer's disease (AD) using the short-form Japanese Verbal Learning Test (JVLT-9). METHODS: Participants were 20 patients with early-stage AD and a control group of 23 healthy older adults (normal controls: NC), each of whom was administered the JVLT-9, which is a verbal list learning task used to assess explicit memory comprehensively. Between-group differences for each score were investigated using the Mann-Whitney U test. A two-way analysis of variance (ANOVA) was performed for the number of correct recalls by group (AD/NC) × JVLT-9 task. In addition, the AD group was divided into a CDR 0.5 group and a CDR 1.0 group, and it was performed as a group (CDR 0.5/1.0) × JVLT-9 task two-way ANOVA. RESULTS: The results demonstrated that the AD group had lower immediate recall, learning rate, semantic clustering, and recognition discrimination and significantly higher intrusion errors compared to the NC group. Further, JVLT-9 recall and recognition rates were found to be lower with higher CDR (an index of dementia severity). CONCLUSION: These results are largely consistent with the features of explicit memory in AD reported in the English version, confirming the clinical utility of the JVLT-9 as a test of explicit memory function.

Stacking Structure of Vaterite Revealed by Atomic Imaging and Diffraction Analysis.

Anhydrous calcium carbonate crystals exist as three polymorphs: calcite, aragonite, and vaterite. Although vaterite is a metastable phase rarely found in the geological environment, it is intriguing that various biominerals are composed of vaterite. The processes of stable vaterite formation in biological systems cannot be understood without elucidating the nature of vaterite. The crystal structure of vaterite has been discussed for nearly a century but is still an open question. Here we propose the actual structure of vaterite by combining atomic imaging and diffraction analysis with simulations of disordered stacking sequences. Vaterite basically appears as layers of hexagonal calcium planes and carbonate (CO3 2-)-containing sheets stacked with +60°, -60°, or 180° rotations from the underlying layer. However, equivalent carbonate positions in alternating layers are forbidden, and four-layer stacking in which the fourth layer rotates 180° relative to the first layer are predominant, forming an orthogonal reciprocal lattice in diffraction patterns. These stacking characteristics replicate the intensity distribution in the electron and X-ray diffraction patterns. This study has almost completely elucidated the crystal structure and stacking sequence of vaterite. Our findings provide insights into the thermodynamic stability of vaterite, which facilitates comprehension of the biomineralization processes and growth dynamics of calcium carbonate.

Acute Bacterial Prostatitis in a 12-Year-Old Boy Without Any Underlying Disease.

Acute bacterial prostatitis (ABP) is a common disease in adults but uncommon in children. Here, we report the case of a pediatric patient without any underlying disease who was diagnosed with ABP while trying to determine the cause of fever refractory to antimicrobial therapy. A previously healthy 12-year-old boy presented with a 13-day history of fever and malaise despite initial antimicrobial treatment. Further tests revealed pyuria and enlarged prostate with possible abscesses, which led to the diagnosis of ABP based on a contrast-enhanced computed tomography (CT) scan. Although initial urine cultures were negative, Corynebacterium pyruviciproducens was detected in subsequent cultures. Antimicrobial therapy for 10 weeks led to improvement without relapse. This case demonstrates that ABP can cause fever in children. Moreover, it shows that contrast-enhanced CT imaging can help identify the cause of fever and that administration of antimicrobials before adequate investigations can confound the diagnosis and complicate the treatment.

Lutein from Chicken Eggs Prevents Amyloid ß-Peptide Aggregation In Vitro and Amyloid ß-Induced Inflammation in Human Macrophages (THP-1).

Epidemiological studies predict that chicken eggs contain constituents other than proteins that prevent Alzheimer's disease. This study screened for constituents that inhibit the aggregation of amyloid ß peptide (Aß)1-42 and elucidated their mechanisms to explore the active components of chicken eggs. Thioflavin T assays and transmission electron microscopy observations showed that arachidonic acid (ARA), lysophosphatidylcholine, lutein (LTN), palmitoleic acid, and zeaxanthin inhibited Aß aggregation. Among these, ARA and LTN showed the highest activity. Photoinduced cross-linking of unmodified protein assays and infrared absorption spectrometry measurements showed that LTN strongly inhibited highly toxic Aß1-42 protofibril formation. Furthermore, LTN suppressed Aß1-42-induced IL 1B and TNF expression in human macrophage-like cells. In summary, LTN plays a crucial role in the AD-preventive effect of chicken eggs by suppressing Aß1-42 aggregation and Aß1-42-induced inflammation.

Diversification of von Willebrand Factor A and Chitin-Binding Domains in Pif/BMSPs Among Mollusks.

Pif is a shell matrix protein (SMP) identified in the nacreous layer of Pinctada fucata (Pfu) comprised two proteins, Pif97 and Pif 80. Pif97 contains a von Willebrand factor A (VWA) and chitin-binding domains, whereas Pif80 can bind calcium carbonate crystals. The VWA domain is conserved in the SMPs of various mollusk species; however, their phylogenetic relationship remains obscure. Furthermore, although the VWA domain participates in protein-protein interactions, its role in shell formation has not been established. Accordingly, in the current study, we investigate the phylogenetic relationship between PfuPif and other VWA domain-containing proteins in major mollusk species. The shell-related proteins containing VWA domains formed a large clade (the Pif/BMSP family) and were classified into eight subfamilies with unique sequential features, expression patterns, and taxa diversity. Furthermore, a pull-down assay using recombinant proteins containing the VWA domain of PfuPif 97 revealed that the VWA domain interacts with five nacreous layer-related SMPs of P. fucata, including Pif 80 and nacrein. Collectively, these results suggest that the VWA domain is important in the formation of organic complexes and participates in shell mineralisation.

Increased brain gyrification and cortical thinning in winter-born patients with schizophrenia spectrum.

Introduction: The findings of epidemiological studies suggest that a relationship exists between the risk of schizophrenia and winter births in the Northern Hemisphere, which may affect the process of fetal neurodevelopment. However, it remains unclear whether birth seasons are associated with the brain morphological characteristics of patients within the schizophrenia spectrum. Methods: The present magnetic resonance imaging study using FreeSurfer software examined the effects of birth seasons (i.e., summer-born vs. winter-born) on the comprehensive brain surface characteristics of 101 patients with schizophrenia (48 summer- and 53 winter-born), 46 with schizotypal disorder (20 summer- and 26 winter-born), and 76 healthy control subjects (28 summer- and 48 winter-born). Results: In comparisons with summer-born patients, winter-born patients, particularly those with schizophrenia, showed significantly increased gyrification mainly in the left lateral occipital and inferior temporal regions and right fronto-parietal region as well as cortical thinning in the right superior frontal region. Birth seasons did not significantly affect the local gyrification index or cortical thickness in healthy controls. Discussion: The present whole-brain surface-based analysis demonstrated that brain morphological characteristics reported in the schizophrenia spectrum were more pronounced in winter-born patients than in summer-born patients, suggesting the contribution of early neurodevelopmental factors associated with birth seasons to the pathophysiology of the schizophrenia spectrum.

Relationships between erythrocyte membrane mono- and poly- unsaturated fatty acid composition and clinical/cognitive indices in antipsychotic-free schizophrenia patients.

Introduction: Membrane phospholipid abnormalities are considered a pathophysiological background for schizophrenia. The aim of the study was to explore in detail the fatty acid (FA) composition in patients with antipsychotic-free schizophrenia and its association with clinical symptoms and cognitive function. Materials and methods: Erythrocyte membrane FAs were measured in 29 antipsychotic-free patients with schizophrenia (male/female = 11/18; mean [standard deviation] age=26.7 [7.9] years) and age and sex-matched 32 healthy volunteers. Clinical symptoms and cognitive function were assessed using the Positive and Negative Syndrome Scale (PANSS), Brief Assessment of Cognition in Schizophrenia (BACS), and the Schizophrenia Cognition Rating Scale (SCoRS). Results: Eicosapentaenoic acid levels were lower in the schizophrenia group than in the healthy control group. In contrast, arachidonic acid and nervonic acid levels were higher in the schizophrenia group than in the control group. Nervonic acid levels were significantly associated with depression scores as measured by the PANSS. No FA levels were correlated with BACS score; however, oleic acid levels were significantly related to cognitive dysfunction, as measured by the SCoRS. Conclusion: These findings suggest that depressive symptoms along with cognitive dysfunction in daily living in schizophrenia may be linked to the FA composition abnormalities. Further studies will be needed to examine potential longitudinal FA changes during the course of schizophrenia as well as disease specificity.

The components of the AhR-molecular chaperone complex differ depending on whether the ligands are toxic or non-toxic.

The aryl hydrocarbon receptor (AhR) forms a complex with the HSP90-XAP2-p23 molecular chaperone when the cells are exposed to toxic compounds. Recently, 1,4-dihydroxy-2-naphthoic acid (DHNA) was reported to be an AhR ligand. Here, we investigated the components of the molecular chaperone complex when DHNA binds to AhR. Proteins eluted from the 3-Methylcolanthrene-affinity column were AhR-HSP90-XAP2-p23 complex. The AhR-molecular chaperone complex did not contain p23 in the eluents from the DHNA-affinity column. In 3-MC-treated cells, AhR formed a complex with HSP90-XAP2-p23 and nuclear translocation occurred within 30 min, while in DHNA-treated cells, AhR formed a complex with AhR-HSP90-XAP2, and translocation was slow from 60 min. Thus, the AhR activation mechanism may differ when DHNA is the ligand compared to toxic ligands.

Organic Matrices of Calcium Carbonate Biominerals Improve Osteoblastic Mineralization.

Many organisms incorporate inorganic solids into their tissues to improve functional and mechanical properties. The resulting mineralized tissues are called biominerals. Several studies have shown that nacreous biominerals induce osteoblastic extracellular mineralization. Among them, Pinctada margaritifera is well known for the ability of its organic matrix to stimulate bone cells. In this context, we aimed to study the effects of shell extracts from three other Pinctada species (Pinctada radiata, Pinctada maxima, and Pinctada fucata) on osteoblastic extracellular matrix mineralization, by using an in vitro model of mouse osteoblastic precursor cells (MC3T3-E1). For a better understanding of the Pinctada-bone mineralization relationship, we evaluated the effects of 4 other nacreous mollusks that are phylogenetically distant and distinct from the Pinctada genus. In addition, we tested 12 non-nacreous mollusks and one extra-group. Biomineral shell powders were prepared, and their organic matrix was partially extracted using ethanol. Firstly, the effect of these powders and extracts was assessed on the viability of MC3T3-E1. Our results indicated that neither the powder nor the ethanol-soluble matrix (ESM) affected cell viability at low concentrations. Then, we evaluated osteoblastic mineralization using Alizarin Red staining and we found a prominent MC3T3-E1 mineralization mainly induced by nacreous biominerals, especially those belonging to the Pinctada genus. However, few non-nacreous biominerals were also able to stimulate the extracellular mineralization. Overall, our findings validate the remarkable ability of CaCO3 biomineral extracts to promote bone mineralization. Nevertheless, further in vitro and in vivo studies are needed to uncover the mechanisms of action of biominerals in bone.

Structural and Functional Analysis of the Amorphous Calcium Carbonate-Binding Protein Paramyosin in the Shell of the Pearl Oyster, Pinctada fucata.

Amorphous calcium carbonate (ACC) is an important precursor phase for the formation of aragonite crystals in the shells of Pinctada fucata. To identify the ACC-binding protein in the inner aragonite layer of the shell, extracts from the shell were used in the ACC-binding experiments. Semiquantitative analyses using liquid chromatography-mass spectrometry revealed that paramyosin was strongly associated with ACC in the shell. We discovered that paramyosin, a major component of the adductor muscle, was included in the myostracum, which is the microstructure of the shell attached to the adductor muscle. Purified paramyosin accumulates calcium carbonate and induces the prism structure of aragonite crystals, which is related to the morphology of prism aragonite crystals in the myostracum. Nuclear magnetic resonance measurements revealed that the Glu-rich region was bound to ACC. Activity of the Glu-rich region was stronger than that of the Asp-rich region. These results suggest that paramyosin in the adductor muscle is involved in the formation of aragonite prisms in the myostracum.

Nomenclature for psychosis risk in Japan: Survey results from high-risk individuals, caregivers, and mental health professionals.

BACKGROUND: Labeling terms for high-risk state for psychosis, such as 'ultra-high risk' (UHR), 'attenuated psychosis syndrome' (APS), and 'at-risk mental state' (ARMS), have been criticized for their potential to lead to stigma. Hence, mental health service users in Melbourne recently proposed new terms illustrating the at-risk concept ['pre-diagnosis stage' (PDS), 'potential of developing a mental illness' (PDMI), and 'disposition for developing a mental illness' (DDMI)]. We aimed at testing the suitability of these existing and new terms in the clinical settings of early psychiatric intervention in Japan. METHODS: At two centers of early intervention (Toyama and Tokyo), a questionnaire on the understanding and opinion of high-risk terminology was administered to 62 high-risk patients, 44 caregivers, and 64 clinicians. The questionnaire contained the existing and new terms, where the term ARMS was translated into two different Japanese terms ARMS-psychosis and ARMS-kokoro. Participants' opinion on the disclosure of high-risk status was also obtained. RESULTS: ARMS-kokoro was most preferred, least stigmatizing, and best explaining the patients' difficulties for all groups, while UHR and other terms including the Japanese word 'psychosis' (i.e., APS and ARMS-psychosis) were not preferred. New labeling terms were generally not well received. All groups preferred full disclosure of high-risk terms by the psychiatrist with or without the presence of family members. CONCLUSION: The term ARMS-kokoro was commonly accepted as a favorable labeling term for the high-risk state for psychosis in Japan. However, another translation ARMS-psychosis was considered stigmatizing, demonstrating the importance of appropriate translation of high-risk terminology into local languages.

Tropomyosin induces the synthesis of magnesian calcite in sea urchin spines.

Calcium carbonate is present in many biominerals, including in the exoskeletons of crustaceans and shells of mollusks. High Mg-containing calcium carbonate was synthesized by high temperatures, high pressures or high molecular organic matter. For example, biogenic high Mg-containing calcite is synthesized under strictly controlled Mg concentration at ambient temperature and pressure. The spines of sea urchins consist of calcite, which contain a high percentage of magnesium. In this study, we investigated the factors that increase the magnesium content in calcite from the spines of the sea urchin, Heliocidaris crassispina. X-ray diffraction and inductively coupled plasma mass spectrometry analyses showed that sea urchin spines contain about 4.8% Mg. The organic matrix extracted from the H. crassispina spines induced the crystallization of amorphous phase and synthesis of magnesium-containing calcite, while amorphous was synthesized without SUE (sea urchin extract). In addition, aragonite was synthesized by SUE treated with protease-K. HC tropomyosin was specifically incorporated into Mg precipitates. Recombinant HC-tropomyosin induced calcite contained 0.1-2.5% Mg synthesis. Western blotting of sea urchin spine extracts confirmed that HC tropomyosin was present in the purple sea urchin spines at a protein weight ratio of 1.5%. These results show that HC tropomyosin is one factor that increases the magnesium concentration in the calcite of H. crassispina spines.

Novel lignin-based extracellular barrier in glandular trichome.

Successful biochemical reactions in organisms necessitate compartmentalization of the requisite components. Glandular trichomes (GTs) act as compartments for the synthesis and storage of specialized compounds. These compounds not only are crucial for the survival of plants under biotic and abiotic stresses but also have medical and commercial value for humans. However, the mechanisms underlying compartmentalization remain unclear. Here we identified a novel structure that is indispensable for the establishment of compartments in cucumber GTs. Silica, a specialized compound, is deposited on the GTs and is visible on the surface of the fruit as a white powder, known as bloom. This deposition provides resistance against pathogens and prevents water loss from the fruits1. Using the cucumber bloomless mutant2, we discovered that a lignin-based cell wall structure in GTs, named 'neck strip', achieves compartmentalization by acting as an extracellular barrier crucial for the silica polymerization. This structure is present in the GTs of diverse plant species. Our findings will enhance the understanding of the biosynthesis of unique compounds in trichomes and provide a basis for improving the production of compounds beneficial to humans.

Refolding, Crystallization, and Crystal Structure Analysis of a Scavenger Receptor Cysteine-Rich Domain of Human Salivary Agglutinin Expressed in Escherichia coli.

Scavenger receptors are a protein superfamily that typically consists of one or more repeats of the scavenger receptor cysteine-rich structural domain (SRCRD), which is an ancient and highly conserved protein module. The expression and purification of eukaryotic proteins containing multiple disulfide bonds has always been challenging. The expression systems that are commonly used to express SRCRD proteins mainly consist of eukaryotic protein expression systems. Herein, we established a high-level expression strategy of a Type B SRCRD unit from human salivary agglutinin using the Escherichia coli expression system, followed by a refolding and purification process. The untagged recombinant SRCRD was expressed in E. coli using the pET-32a vector, which was followed by a refolding process using the GSH/GSSG redox system. The SRCRD expressed in E. coli SHuffle T7 showed better solubility after refolding than that expressed in E. coli BL21(DE3), suggesting the importance of the disulfide bond content prior to refolding. The quality of the refolded protein was finally assessed using crystallization and crystal structure analysis. As proteins refolded from inclusion bodies exhibit a high crystal quality and reproducibility, this method is considered a reliable strategy for SRCRD protein expression and purification. To further confirm the structural integrity of the refolded SRCRD protein, the purified protein was subjected to crystallization using sitting-drop vapor diffusion method. The obtained crystals of SRCRD diffracted X-rays to a resolution of 1.47 Å. The solved crystal structure appeared to be highly conserved, with four disulfide bonds appropriately formed. The surface charge distribution of homologous SRCRD proteins indicates that the negatively charged region at the surface is associated with their calcium-dependent ligand recognition. These results suggest that a high-quality SRCRD protein expressed by E. coli SHuffle T7 can be successfully folded and purified, providing new options for the expression of members of the scavenger receptor superfamily.

The relationship between gray/white matter contrast and cognitive performance in first-episode schizophrenia.

Previous postmortem brain studies have revealed disturbed myelination in the intracortical regions in patients with schizophrenia, possibly reflecting anomalous brain maturational processes. However, it currently remains unclear whether this anomalous myelination is already present in early illness stages and/or progresses during the course of the illness. In this magnetic resonance imaging study, we examined gray/white matter contrast (GWC) as a potential marker of intracortical myelination in 63 first-episode schizophrenia (FESz) patients and 77 healthy controls (HC). Furthermore, we investigated the relationships between GWC findings and clinical/cognitive variables in FESz patients. GWC in the bilateral temporal, parietal, occipital, and insular regions was significantly higher in FESz patients than in HC, which was partly associated with the durations of illness and medication, the onset age, and lower executive and verbal learning performances. Because higher GWC implicates lower myelin in the deeper layers of the cortex, these results suggest that schizophrenia patients have less intracortical myelin at the time of their first psychotic episode, which underlies lower cognitive performance in early illness stages.

Atmospheric CO2 Sequestration in Seawater Enhanced by Molluscan Shell Powders.

Carbon capture, utilization, and storage (CCUS) are widely recognized as a promising technology for mitigating climate change. CO2 mineralization using Ca-rich fluids and high-concentration CO2 gas has been studied extensively. However, few studies have reported CO2 mineralization with atmospheric CO2, owing to the difficulty associated with its low concentration. In seawater, the biomineralization process promotes Ca accumulation and CaCO3 precipitation, assisted by specific organic matter. In this study, we examined the conversion of atmospheric CO2 into CaCO3 in seawater using shell powders (Pinctada fucata, Haliotis discus, Crassostrea gigas, Mizuhopecten yessoensis, Turbo sazae, and Saxidomus purpurata). Among the six species, the shell powder of S. purpurata showed the highest rate of CaCO3 formation and recovery of CaCO3. NaClO treatment test revealed that the organic matter in the shells enhanced the CO2 mineralization. All materials used in this study, including atmospheric CO2, seawater, and shells, are economically feasible for large-scale applications. Using shell powder for CO2 mineralization in seawater embodies an innovative technological advancement to address climate change.

Association between copy number variations in parkin (PRKN) and schizophrenia and autism spectrum disorder: A case-control study.

AIM: The present study aimed to examine the association between copy number variations (CNVs) in parkin (PRKN) and schizophrenia (SCZ) and autism spectrum disorder (ASD) in a large case-control sample. METHOD: Array comparative genomic hybridization was performed on 3111 cases with SCZ, 1236 cases with ASD, and 2713 controls. We systematically prioritized likely pathogenic CNVs (LP-CNVs) in PRKN and examined their association with SCZ and ASD. RESULTS: In total, 3014 SCZ cases (96.9%), 1205 ASD cases (97.5%), and 2671 controls (98.5%) passed quality control. We found that monoallelic carriers of LP-CNVs in PRKN were common (70/6890, 1.02%) and were not at higher risk of SCZ (p = 0.29) or ASD (p = 0.72). We observed that the distribution pattern of LP-CNVs in the Japanese population was consistent with those in other populations. We also identified a patient diagnosed with SCZ and early-onset Parkinson's disease carrying biallelic pathogenic CNVs in PRKN. The absence of Parkinson's symptoms in 10 other monoallelic carriers of the same pathogenic CNV further reflects the lack of effect of monoallelic pathogenic variants in PRKN in the absence of a second hit. CONCLUSION: The present findings suggest that monoallelic CNVs in PRKN do not confer a significant risk for SCZ or ASD. However, further studies to investigate the association between biallelic CNVs in PRKN and SCZ and ASD are warranted.

Molecular mechanism of Fe3+ binding inhibition to Vibrio metschnikovii ferric ion-binding protein, FbpA, by rosmarinic acid and its hydrolysate, danshensu.

Global warming has increased the growth of pathogenic Vibrio bacteria, which can cause foodborne illnesses and death. Vibrio bacteria require iron for growth and survival. They utilize a ferric ion-binding protein (FbpA) to bind and transport Fe3+ into the cell. FbpA from Vibrio metschnikovii (Vm) is a potential target for inhibiting its growth. Rosmarinic acid (RA) can block the binding of VmFbpA to Fe3+ ; however, the molecular mechanism of Fe3+ binding and RA inhibition to VmFbpA is unclear. In this study, we used x-ray crystallography to determine the Fe3+ -binding mode of VmFbpA and the mechanism of RA inhibition. The structures revealed that in the Fe3+ bound form, Fe3+ was coordinated to VmFbpA by two Tyr residues, two HCO3 - ions, and two water molecules in a six-coordinated geometry. In contrast, in the inhibitor bound form, RA was initially bound to VmFbpA following gel filtration purification, but it was hydrolyzed to danshensu (DSS), which occupied the binding site as shown in an electron density map and reverse phase chromatography (RPC) analysis. Both RA and DSS exhibited a stronger binding affinity to VmFbpA, higher Fe3+ reduction capacity, and more potent bacteriostatic effect on V. metschnikovii compared with caffeic acid (CA), another hydrolysis product of RA. These results provide insight into the mechanism of iron acquisition by V. metschnikovii and inhibition by RA and DSS. Our findings offer clues on the development of effective strategies to prevent Vibrio infections.

A Case of Uncomplicated Bacteremia Caused by Capnocytophaga canimorsus in an Immunocompetent Patient.

This report describes uncomplicated bacteremia caused by Capnocytophaga canimorsus in an immunocompetent woman who presented with rigor and fever. She was hemodynamically stable. Two blood samples were immediately cultured because rigor indicated bacteremia. Although her symptoms were relieved, Gram-negative rods grew from blood cultures. She noted that she had been bitten by her dog before the first examination. The bacterium was confirmed as C. canimorsus by gene analysis. Infection with C. canimorsus can be fatal when accompanied by sepsis in elderly or immunocompromised patients. However, this case was considered rare as the patient was 41 years old and immunocompetent.

Comparative genomic and transcriptomic analyses of African swine fever virus strains.

African swine fever (ASF) is the most devastating disease caused by the African swine fever virus (ASFV), impacting the pig industry worldwide and threatening food security and biodiversity. Although two vaccines have been approved in Vietnam to combat ASFV, the complexity of the virus, with its numerous open reading frames (ORFs), necessitates a more diverse vaccine strategy. Therefore, we focused on identifying and investigating the potential vaccine targets for developing a broad-spectrum defense against the virus. This study collected the genomic and/or transcriptomic data of different ASFV strains, specifically from in vitro studies, focusing on comparisons between genotypes I, II, and X, from the National Center for Biotechnology Information (NCBI) database. The comprehensive analysis of the genomic and transcriptomic differences between high- and low-virulence strains revealed six early genes, 13 late genes, and six short genes as potentially essential ORFs associated with high-virulence. In addition, many other ORFs (e.g., 14 multigene family members) are worth investigating. The results of this study provided candidate ORFs for developing ASF vaccines and therapies.