A highly efficient blocking ELISA based on p72 monoclonal antibody for the detection of African swine fever virus antibodies and identification of its linear B cell epitope.

Due to the absence of effective vaccine and treatment, African swine fever virus (ASFV) control is entirely dependent on accurate and early diagnosis, along with culling of infected pigs. The B646L/p72 is the major capsid protein of ASFV and is an important target for developing a diagnostic assays and vaccines. Herein, we generated a monoclonal antibody (mAb) (designated as 2F11) against the trimeric p72 protein, and a blocking ELISA (bELISA) was established for the detection of both genotype I and II ASFV antibodies. To evaluate the performance of the diagnostic test, a total of 506 porcine serum samples were tested. The average value of percent of inhibition (PI) of 133 negative pig serum was 8.4 % with standard deviation (SD) 6.5 %. Accordingly, the cut-off value of the newly established method was set at 28 % (mean + 3SD). Similarly, a receiver operating characteristic (ROC) was applied to determine the cut off value and the p72-bELISA exhibited a sensitivity of 100 % and a specificity of 99.33 % when the detection threshold was set at 28 %. The bELISA was also able to specifically recognize anti-ASFV sera without cross-reacting with other positive serums for other major swine pathogens. Moreover, by designing a series of overlapped p72 truncated proteins, the linear B cell epitope recognized by 2F11 mAb was defined to be 283NSHNIQ288. Amino acid sequence comparison revealed that the amino acid sequence 283NSHNIQ288 is highly conserved between different ASFV isolates. Our findings indicate that the newly established mAb based blocking ELISA may have a great potential in improving the detection of ASFV antibodies and provides solid foundation for further studies.

Multiplexed CRISPR-Based Nucleic Acid Detection Using a Single Cas Protein.

Thanks to its ease, speed, and sensitivity, CRISPR-based nucleic acid detection has been increasingly explored for molecular diagnostics. However, one of its major limitations is lack of multiplexing capability because the detection relies on the trans-cleavage activity of the Cas protein, which necessitates the use of multiple orthogonal Cas proteins for multiplex detection. Here we report the development of a multiplexed CRISPR-based nucleic acid detection system with single-nucleotide resolution using a single Cas protein (Cas12a). This method, termed as CRISPR-TMSD, integrates the toehold-mediated strand displacement (TMSD) reaction, and the cis-cleavage activity of the Cas protein and multiplexed detection are achieved using a single Cas protein owing to the use of target-specific reporters. A set of computational simulation toolkits was used to design the TMSD reporter, allowing for highly sensitive and specific identification of target sequences. In combination with the recombinase polymerase amplification (RPA), the detection limit can reach as low as 1 copy/µL. As proof of concept, CRISPR-TMSD was subsequently used to detect an oncogenic gene and SARS-CoV-2 RNA with a single-nucleotide resolution. This work represents a conceptually new strategy for designing a CRISPR-based diagnostic system and has great potential to expand the application of CRISPR-based diagnostics.

Applying the digital data and the bioinformatics tools in SARS-CoV-2 research.

Bioinformatics has been playing a crucial role in the scientific progress to fight against the pandemic of the coronavirus disease 2019 (COVID-19) caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). The advances in novel algorithms, mega data technology, artificial intelligence and deep learning assisted the development of novel bioinformatics tools to analyze daily increasing SARS-CoV-2 data in the past years. These tools were applied in genomic analyses, evolutionary tracking, epidemiological analyses, protein structure interpretation, studies in virus-host interaction and clinical performance. To promote the in-silico analysis in the future, we conducted a review which summarized the databases, web services and software applied in SARS-CoV-2 research. Those digital resources applied in SARS-CoV-2 research may also potentially contribute to the research in other coronavirus and non-coronavirus viruses.

ERK and USP5 govern PD-1 homeostasis via deubiquitination to modulate tumor immunotherapy.

The programmed cell death protein 1 (PD-1) is an inhibitory receptor on T cells and plays an important role in promoting cancer immune evasion. While ubiquitin E3 ligases regulating PD-1 stability have been reported, deubiquitinases governing PD-1 homeostasis to modulate tumor immunotherapy remain unknown. Here, we identify the ubiquitin-specific protease 5 (USP5) as a bona fide deubiquitinase for PD-1. Mechanistically, USP5 interacts with PD-1, leading to deubiquitination and stabilization of PD-1. Moreover, extracellular signal-regulated kinase (ERK) phosphorylates PD-1 at Thr234 and promotes PD-1 interaction with USP5. Conditional knockout of Usp5 in T cells increases the production of effector cytokines and retards tumor growth in mice. USP5 inhibition in combination with Trametinib or anti-CTLA-4 has an additive effect on suppressing tumor growth in mice. Together, this study describes a molecular mechanism of ERK/USP5-mediated regulation of PD-1 and identifies potential combinatorial therapeutic strategies for enhancing anti-tumor efficacy.

Protein engineering of invertase for enhancing yeast dough fermentation under high-sucrose conditions.

During yeast dough fermentation, such as the high-sucrose bread-making process, the yeast cells are subjected to considerable osmotic stress, resulting in poor outcomes. Invertase is important for catalyzing the irreversible hydrolysis of sucrose to free glucose and fructose, and decreasing the catalytic activity of the invertase may reduce the glucose osmotic stress on the yeast. In this study, we performed structural design and site-directed mutagenesis (SDM) on the Saccharomyces cerevisiae invertase (ScInV) in an Escherichia coli expression system to study the catalytic activity of ScInV mutants in vitro. In addition, we generated the same mutation sites in the yeast endogenous genome and tested their invertase activity in yeast and dough fermentation ability. Our results indicated that appropriately reduced invertase activity of yeast ScInV can enhance dough fermentation activity under high-sucrose conditions by 52%. Our systems have greatly accelerated the engineering of yeast endogenous enzymes both in vitro and in yeast, and shed light on future metabolic engineering of yeast.

Comparative Physiological and Transcriptomic Analyses of Two Contrasting Pepper Genotypes under Salt Stress Reveal Complex Salt Tolerance Mechanisms in Seedlings.

As a glycophyte plant, pepper (Capsicum annuum L.) is widely cultivated worldwide, but its growth is susceptible to salinity damage, especially at the seedling stage. Here, we conducted a study to determine the physiological and transcriptional differences between two genotype seedlings (P300 and 323F3) with contrasting tolerance under salt stress. The P300 seedlings were more salt-tolerant and had higher K+ contents, higher antioxidase activities, higher compatible solutes, and lower Na+ contents in both their roots and their leaves than the 323F3 seedlings. During RNA-seq analysis of the roots, more up-regulated genes and fewer down-regulated genes were identified between salt-treated P300 seedlings and the controls than between salt-treated 323F3 and the controls. Many ROS-scavenging genes and several SOS pathway genes were significantly induced by salt stress and exhibited higher expressions in the salt-treated roots of the P300 seedlings than those of 323F3 seedlings. Moreover, biosynthesis of the unsaturated fatty acids pathway and protein processing in the endoplasmic reticulum pathway were deeply involved in the responses of P300 to salt stress, and most of the differentially expressed genes involved in the two pathways, including the genes that encode mega-6 fatty acid desaturases and heat-shock proteins, were up-regulated. We also found differences in the hormone synthesis and signaling pathway genes in both the P300 and 323F3 varieties under salt stress. Overall, our results provide valuable insights into the physiological and molecular mechanisms that affect the salt tolerance of pepper seedlings, and present some candidate genes for improving salt tolerance in pepper.

Structural Design and Assessing of Recombinantly Expressed African Swine Fever Virus p72 Trimer in Saccharomyces cerevisiae.

In an effort to control the outbreak of the African Swine Fever Virus (ASFV), there is an urgent need to develop an effective method to prevent the pandemic, including vaccines and diagnostic methods. The major capsid protein of ASFV p72 (B646L), which forms a trimer with each monomer adopting a double jelly roll fold, is the main component of the virus particle and major antigen of ASFV. Thus, the p72 protein may be considered an antigen candidate for vaccine and diagnostic development. However, the development of ASFV p72 trimer for the industry application, including veterinary usage, faces unavoidable challenges: firstly, the low cost of the antigen production is required in vaccine and diagnostic application; and, secondly, whether produced antigen folds in its native conformation. Here, based on the information provided by the atomic structure of p72, we have successfully performed rational mutagenesis on p72 trimers and expressed it in Saccharomyces cerevisiae with high yields. The cryo-EM structure of recombinant expressed p72 trimer is determined at 4.18 Å in resolution. The correlation coefficient between this structure and the ASFV virus structure is 0.77, suggesting a highly similar fold of this trimer with the native protein on the virus particle.

Structure of Senecavirus A 3C Protease Revealed the Cleavage Pattern of 3C Protease in Picornaviruses.

Senecavirus A (SVA) is an emerging picornavirus infecting porcine of all age groups and causing foot and mouth disease (FMD)-like symptoms. One of its key enzymes is the 3C protease (3Cpro), which is similar to other picornaviruses and essential for virus maturation by controlling polyprotein cleavage and RNA replication. In this study, we reported the crystal structure of SVA 3Cpro at a resolution of 1.9 Å and a thorough structural comparison against all published picornavirus 3Cpro structures. Using statistical and graphical visualization techniques, we also investigated the sequence specificity of the 3Cpro. The structure revealed that SVA 3Cpro adopted a typical chymotrypsin-like fold with the S1 subsite as the most conservative site among picornavirus 3Cpro. The surface loop, A1-B1 hairpin, adopted a novel conformation in SVA 3Cpro and formed a positively charged protrusion around S' subsites. Correspondingly, SVA scissile bonds preferred Asp rather than neutral amino acids at P3' and P4'. Moreover, SVA 3Cpro showed a wide range tolerance to P4 residue volume (acceptable range: 67 Å3 to 141 Å3), such as aromatic side chain, in contrast to other picornaviruses. In summary, our results provided valuable information for understanding the cleavage pattern of 3Cpro. IMPORTANCE Picornaviridae is a group of RNA viruses that harm both humans and livestock. 3Cpro is an essential enzyme for picornavirus maturation, which makes it a promising target for antiviral drug development and a critical component for virus-like particle (VLP) production. However, the current challenge in the development of antiviral drugs and VLP vaccines includes the limited knowledge of how subsite structure determines the 3Cpro cleavage pattern. Thus, an extensive comparative study of various picornaviral 3Cpro was required. Here, we showed the 1.9 Å crystal structure of SVA 3Cpro. The structure revealed similarities and differences in the substrate-binding groove among picornaviruses, providing new insights into the development of inhibitors and VLP.

Purification-induced damage to calicivirus particles at near-atomic resolution.

The purification of virus particles is an essential process for the manufacture of vaccines. However, the application of different purification processes may affect the quality of the virus particles, such as structural integrity and homogeneity, which may further influence the infectivity and immunogenicity of the purified virus. In this study, we took Feline calicivirus (FCV), a common natural pathogen in cats belonging to Caliciviridae, as a research model. By using cryo-electron microscopy (cryo-EM), we incorporated the 3D classification process as a virus flexibility evaluation system. Cryo-EM images of virus particles resulting from different purification processes were compared at near-atomic resolution. The results indicated that molecular sieving purification will impact the stability of P-domains through increasing flexibility as determined by the evaluation system, which can be extended to assess the purification effect on the entire particle. This evaluation process can be further applied to all non-enveloped viruses.

Melatonin Alleviates Copper Toxicity via Improving ROS Metabolism and Antioxidant Defense Response in Tomato Seedlings.

The excessive accumulation of copper (Cu2+) has become a threat to worldwide crop production. Recently, it was revealed that melatonin (MT) could play a crucial role against heavy metal (HM) stresses in plants. However, the underlying mechanism of MT function acted upon by Cu2+ stress (CS) has not been substantiated in tomatoes. In the present work, we produced MT-rich tomato plants by foliar usage of MT, and MT-deficient tomato plants by employing a virus-induced gene silencing methodology and exogenous foliar application of MT synthesis inhibitor para-chlorophenylalanine (pCPA). The obtained results indicate that exogenous MT meaningfully alleviated the dwarf phenotype and impeded the reduction in plant growth caused by excess Cu2+. Furthermore, MT effectively restricted the generation of reactive oxygen species (ROS) and habilitated cellular integrity by triggering antioxidant enzyme activities, especially via CAT and APX, but not SOD and POD. In addition, MT increased nonenzymatic antioxidant activity, including FRAP and the GSH/GSSG and ASA/DHA ratios. MT usage improved the expression of several defense genes (CAT, APX, GR and MDHAR) and MT biosynthesis-related genes (TDC, SNAT and COMT). Taken together, our results preliminarily reveal that MT alleviates Cu2+ toxicity via ROS scavenging, enhancing antioxidant capacity when subjected to excessive Cu2+. These results build a solid foundation for developing new insights to solve problems related to CS.

Molecular breeding of barley for quality traits and resilience to climate change.

Barley grains are a rich source of compounds, such as resistant starch, beta-glucans and anthocyanins, that can be explored in order to develop various products to support human health, while lignocellulose in straw can be optimised for feed in husbandry, bioconversion into bioethanol or as a starting material for new compounds. Existing natural variations of these compounds can be used to breed improved cultivars or integrated with a large number of mutant lines. The technical demands can be in opposition depending on barley's end use as feed or food or as a source of biofuel. For example beta-glucans are beneficial in human diets but can lead to issues in brewing and poultry feed. Barley breeders have taken action to integrate new technologies, such as induced mutations, transgenics, marker-assisted selection, genomic selection, site-directed mutagenesis and lastly machine learning, in order to improve quality traits. Although only a limited number of cultivars with new quality traits have so far reached the market, research has provided valuable knowledge and inspiration for future design and a combination of methodologies to achieve the desired traits. The changes in climate is expected to affect the quality of the harvested grain and it is already a challenge to mitigate the unpredictable seasonal and annual variations in temperature and precipitation under elevated [CO2] by breeding. This paper presents the mutants and encoded proteins, with a particular focus on anthocyanins and lignocellulose, that have been identified and characterised in detail and can provide inspiration for continued breeding to achieve desired grain and straw qualities.

Correlation analysis of plasma microRNA-122 and microRNA-33a with severity of coronary artery disease in patients with type 2 diabetes mellitus and coronary heart disease / 中国医师进修杂志

Objective:To investigate the correlation between plasma microRNA (miR)-122, miR-33a and the severity of coronary artery disease in patients with type 2 diabetes mellitus (T2DM) and coronary heart disease.Methods:The clinical data of 196 patients with T2DM from January 2019 to October 2021 in Xuzhou First People′s Hospital were retrospectively analyzed. Among them, 81 cases were complicated with coronary heart disease (combined group), 115 cases were not complicated with coronary heart disease (control group). The plasma levels of miR-122 and miR-33a were detected by real-time fluorescence quantitative reverse transcription polymerase chain reaction, the plasma level of N-terminal B-type natriuretic peptide precursor (NT-proBNP) was detected by enzyme-linked immunosorbent assay. In combined group, the number of coronary artery lesions was determined according to the results of coronary angiography, and Gensini score was evaluated. Linear regression model was used to analyze the relationship between plasma miR-122, miR-33a and NT-proBNP levels with the incidence of coronary heart disease in patients with T2DM. Receiver operating characteristic (ROC) curve was used to analyze the plasma miR-122 and miR-33a in predicting efficiency of coronary heart disease in patients with T2DM. In combined group, Spearman correlation method was used to analyze the relationship between plasma miR-122, miR-33a and the number of coronary artery lesions, and Pearson correlation method was used to analyze the relationship between plasma miR-122, miR-33a and plasma NT proBNP, Gensini score.Results:The plasma miR-122, miR-33a and NT-proBNP in combined group were significantly higher than those in control group: 5.76 ± 1.35 vs. 1.18 ± 0.33, 1.39 ± 0.37 vs. 0.65 ± 0.11 and (786.87 ± 156.39) ng/L vs. (103.45 ± 19.27) ng/L respectively, and there were statistical differences ( P<0.01). Linear regression result showed that plasma miR-122, miR-33a, and NT-proBNP were positive correlation with occurrence of coronary heart disease in patients with T2DM ( P<0.01); ROC curve analysis result showed that the area under curve of plasma miR-122, miR-33a and combination in predicting coronary heart disease in patients with T2DM were 0.816, 0.845 and 0.912 respectively (95% CI 0.744 to 0.865, 0.768 to 0.892 and 0.836 to 0.967). Coronary angiography result showed that there were 46 cases of single vessel lesions, 25 cases of double vessel lesions and 10 cases of three vessel lesions. The plasma miR-122, miR-33a, NT-proBNP and Gensini score in patients with three vessel lesions were significantly higher than those in patients with double vessel lesions and patients with single vessel lesions: 6.52 ± 0.96 vs. 4.95 ± 0.85 and 3.74 ± 0.52, 1.45 ± 0.31 vs. 1.06 ± 0.25 and 0.81 ± 0.13, (829.78 ± 62.59) ng/L vs. (627.48 ± 47.12) and (502.64 ± 38.24) ng/L, (63.89 ± 12.71) scores vs. (42.18 ± 6.03) and (22.36 ± 2.41) scores, the indexes in patients with double vessel lesions were significantly higher than those in patients with single vessel lesions, and there were statistical differences ( P<0.05). In combined group, Spearman correlation analysis result showed that the plasma miR-122 and miR-33a were positive correlation with the number of coronary artery lesions ( r = 0.879 and 0.825, P<0.05); Pearson correlation analysis result showed that the plasma miR-122 and miR-33a were positive correlation with the plasma NT-proBNP and Gensini score (miR-122: r = 0.896 and 0.788, miR-33a: r = 0.871 and 0.765; P<0.05). Conclusions:The plasma levels of miR-122 and miR-33a are related to the occurrence of coronary heart disease and severity of coronary artery disease in patients with T2DM, which may be used to guide the prevention and treatment of coronary heart disease in patients with T2DM.

Nucleocapsid mutations R203K/G204R increase the infectivity, fitness, and virulence of SARS-CoV-2.

Previous work found that the co-occurring mutations R203K/G204R on the SARS-CoV-2 nucleocapsid (N) protein are increasing in frequency among emerging variants of concern or interest. Through a combination of in silico analyses, this study demonstrates that R203K/G204R are adaptive, while large-scale phylogenetic analyses indicate that R203K/G204R associate with the emergence of the high-transmissibility SARS-CoV-2 lineage B.1.1.7. Competition experiments suggest that the 203K/204R variants possess a replication advantage over the preceding R203/G204 variants, possibly related to ribonucleocapsid (RNP) assembly. Moreover, the 203K/204R virus shows increased infectivity in human lung cells and hamsters. Accordingly, we observe a positive association between increased COVID-19 severity and sample frequency of 203K/204R. Our work suggests that the 203K/204R mutations contribute to the increased transmission and virulence of select SARS-CoV-2 variants. In addition to mutations in the spike protein, mutations in the nucleocapsid protein are important for viral spreading during the pandemic.

Characterisation of the class III peroxidase gene family in carrot taproots and its role in anthocyanin and lignin accumulation.

Plant class III peroxidases (CIII Prxs) are involved in numerous essential plant life processes, such as plant development and differentiation, lignification and seed germination, and defence against pathogens. However, there is limited information about the structure-function relationships of Prxs in carrots. This study identified 75 carrot peroxidases (DcPrxs) and classified them into seven subgroups based on phylogenetic analysis. Gene structure analysis revealed that these DcPrxs had between one and eight introns, while conserved motif analysis showed a typical motif composition and arrangement for CIII Prx. In addition, eighteen tandem duplication events, but only eight segmental duplications, were identified among these DcPrxs, indicating that tandem duplication was the main contributor to the expansion of this gene family. Histochemical analyses showed that lignin was mainly localised in the cell walls of xylem, and Prx activity was determined in the epidermal region of taproots. The xylem always showed higher lignin concentration and lower Prx activity compared to the phloem in the taproots of both carrot cultivars. Combining these observations with RNA sequencing, some Prx genes were identified as candidate genes related to lignification and pigmentation. Three peroxidases (DcPrx30, DcPrx32, DcPrx62) were upregulated in the phloem of both genotypes. Carrot taproots are an attractive resource for natural food colourants and this study elucidated genome-wide insights of Prx for the first time, developing hypotheses concerning their involvement with lignin and anthocyanin in purple carrots. The findings provide an essential foundation for further studies of Prx genes in carrot, especially on pigmentation and lignification mechanisms.

Rapid Golden Gate assembly of exons from genomic DNA for protein expression in Escherichia coli and Pichia pastoris.

The development of a quick, single-step cloning system for generation of multiexon gene expression constructs is presented. The system allows efficient and cost-effective assembly of multiple exons of interest genes into different expression plasmids in both Escherichia coli and Pichia pastoris. The high cloning efficiency and low cost of the system make it ideal for a novel workflow for the assembly of intron-bearing genes for expression in two different expression hosts.

Rapid and sensitive RPA-Cas12a-fluorescence assay for point-of-care detection of African swine fever virus.

African swine fever (ASF) is a serious contagious disease that causes fatal haemorrhagic fever in domestic and wild pigs, with high morbidity. It has caused devastating damage to the swine industry worldwide, necessitating the focus of attention on detection of the ASF pathogen, the African swine fever virus (ASFV). In order to overcome the disadvantages of conventional diagnostic methods (e.g. time-consuming, demanding and unintuitive), quick detection tools with higher sensitivity need to be explored. In this study, based on the conserved p72 gene sequence of ASFV, we combined the Cas12a-based assay with recombinase polymerase amplification (RPA) and a fluorophore-quencher (FQ)-labeled reporter assay for rapid and visible detection. Five crRNAs designed for Cas12a-based assay showed specificity with remarkable fluorescence intensity under visual inspection. Within 20 minutes, with an initial concentration of two copies of DNA, the assay can produce significant differences between experimental and negative groups, indicating the high sensitivity and rapidity of the method. Overall, the developed RPA-Cas12a-fluorescence assay provides a fast and visible tool for point-of-care ASFV detection with high sensitivity and specificity, which can be rapidly performed on-site under isothermal conditions, promising better control and prevention of ASF.

A CRISPR-Cas9 based shuffle system for endogenous histone H3 and H4 combinatorial mutagenesis.

Post-translational modifications of histone proteins greatly impact gene expression and cell fate decisions in eukaryotes. To study these, it is important to develop a convenient, multiplex, and efficient method to precisely introduce mutations to histones. Because eukaryotic cells usually contain multiple copies of histone genes, it is a challenge to mutate all histones at the same time by the traditional homologous recombination method. Here, we developed a CRISPR-Cas9 based shuffle system in Saccharomyces cerevisiae, to generate point mutations on both endogenous histone H3 and H4 genes in a rapid, seamless and multiplex fashion. Using this method, we generated yeast strains containing histone triple H3-K4R-K36R-K79R mutants and histone combinatorial H3-K56Q-H4-K59A double mutants with high efficiencies (70-80%). This CRISPR-Cas9 based mutagenesis system could be an invaluable tool to the epigenetics field.

Rapid Spread of Mutant Alleles in Worldwide SARS-CoV-2 Strains Revealed by Genome-Wide Single Nucleotide Polymorphism and Variation Analysis.

The novel coronavirus (SARS-CoV-2) has become a pandemic and is threatening human health globally. Here, we report nine newly evolved SARS-CoV-2 single nucleotide polymorphism (SNP) alleles those underwent a rapid increase (seven cases) or decrease (two cases) in their frequency for 30-80% in the initial four months, which are further confirmed by intrahost single nucleotide variation analysis using raw sequence data including 8,217 samples. The nine SNPs are mostly (8/9) located in the coding region and are mainly (6/9) nonsynonymous substitutions. The nine SNPs show a complete linkage in SNP pairs and belong to three different linkage groups, named LG_1 to LG_3. Analyses in population genetics show signatures of adaptive selection toward the mutants in LG_1, but no signal of selection for LG_2. Population genetic analysis results on LG_3 show geological differentiation. Analyses on geographic COVID-19 cases and published clinical data provide evidence that the mutants in LG_1 and LG_3 benefit virus replication and those in LG_1 have a positive correlation with the disease severity in COVID-19-infected patients. The mutants in LG_2 show a bias toward mildness of the disease based on available public clinical data. Our findings may be instructive for epidemiological surveys and disease control of COVID-19 in the future.

Lateral Flow Assay for the Detection of African Swine Fever Virus Antibodies Using Gold Nanoparticle-Labeled Acid-Treated p72.

African swine fever is a widespread and highly contagious disease in the porcine population, which is caused by African swine fever virus (ASFV). The PCR and ELISA detection methods are the main conventional diagnostic methods for ASFV antigen/antibody detection in the field. However, these methods have limitations of expensive equipment, trained technicians, and time-consuming results. Thus, a rapid, inexpensive, accurate and on-site detection method is urgently needed. Here we describe a double-antigen-sandwich lateral-flow assay based on gold nanoparticle-conjugated ASFV major capsid protein p72, which can detect ASFV antibody in serum samples with high sensitivity and specificity in 10 min and the results can be determined by naked eyes. A lateral flow assay was established by using yeast-expressed and acid-treated ASFV p72 conjugated with gold nanoparticles, which are synthesized by seeding method. A high coincidence (97.8%) of the assay was determined using clinical serum compared to a commercial ELISA kit. In addition, our lateral flow strip can detect as far as 1:10,000 diluted clinically positive serum for demonstration of high sensitivity. In summary, the assay developed here was shown to be rapid, inexpensive, accurate and highly selective. It represents a reliable method for on-site ASFV antibody detection and may help to control the ASFV pandemic.