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A smartphone-integrated plasmonic biosensor for amplification-free detection of African swine fever virus.

African Swine Fever Virus (ASFV) poses a catastrophic threat to global swine production, with recent outbreaks across Europe, Asia, and the Caribbean, significantly elevating the biosecurity risk to the United States' billion-dollar pork industry. Current diagnostic gold standards are laboratory-dependent and introduce critical delays in outbreak response. To address this gap, a plasmonic biosensor based on functionalized gold nanoparticles (GNPs) was developed for the rapid, amplification-free detection of ASFV. GNPs were surface-functionalized with 11-mercaptoundecanoic acid (MUDA) and combined in situ with ASFV-specific oligonucleotide probes targeting a conserved region of the p72 (B646L) gene. The detection mechanism relies on acid-induced aggregation: hybridization of target ASFV DNA to the probe generates a rigid duplex that shields the nanoparticles from acid-induced destabilization, maintaining a ruby-red color, whereas in the absence of target DNA the GNPs aggregate, producing a visible red-to-blue color shift. The optimized plasmonic biosensor demonstrated 100% analytical specificity, with no cross-reactivity against a panel of 19 non-target bacterial genomic DNA samples representative of the swine environment. Detection limits determined by the IUPAC 3σ criterion were 285 copies per reaction for Probe 1 and 402 copies per reaction for Probe 2, within the same order of magnitude as the qPCR reference assay run on the same dilution series (approximately 312 copies per reaction) under the experimental conditions used here. A smartphone-based Bio-Analytics App employing an RGB color-conversion algorithm served as a quantitative reader, yielding signal-to-noise ratios (S/N) that strongly correlated with benchtop spectrophotometric readings (A520/A620 ratio, R2 = 0.96) and achieved diagnostic concordance with qPCR binary calls. This platform offers a robust and low-cost (∼$2 per test), amplification-free approach to ASFV screening with potential for point-of-need deployment, subject to future validation in clinical specimens.

Journal Article

'PePApipe': A complete bioinformatics analysis pipeline for African Swine Fever Virus genome.

African Swine Fever Virus (ASFV) is of high concern in porcine livestock across the world due to both the high mortality rates and the trade restrictions imposed on affected regions. The viral genome is large and complex, and genomic analysis is essential for tracing its origin and evolution. Although several bioinformatics tools exist for genome assembly and analysis, no single platform integrates all necessary steps in an accessible and systematic way. In this study the authors developed 'PePApipe', a custom-built, user-friendly pipeline that enables rapid, complete, and efficient ASFV genome analysis. It is specifically designed for laboratory professionals with limited bioinformatics experience, requiring only basic command-line knowledge. Starting from raw sequencing data, PePApipe integrates thirteen software tools into one automated workflow, covering quality control and pre-processing of raw reads, de novo genome assembly and variant calling. Programmed in Python, it can be executed locally through bash scripts, or using a Slurm protocol for batch processing of multiple samples. The main outputs are the ASFV consensus genome sequence and a file listing its putative variants compared to the selected reference genome. PePApipe classifies generated files into structured folders and produces intermediate files that can be used as inputs for further or parallel analyses; users can also enable or disable specific steps in each particular case. This pipeline is adaptable and complementary to downstream steps such as viral genome annotation or genome visualization. By consolidating all stages of viral genome analysis into a single automated workflow, PePApipe reduces the likelihood of user error, and enhances reproducibility and efficiency. This user-friendly pipeline facilitates the transition from sequencing to assembly and downstream analysis of viral genomes, ensuring a fast and reliable response to molecular analysis demands. Finally, the pipeline can be easily adapted to the study of other viral species, expanding its application in infectious diseases surveillance.

African Swine Fever Virus

Cross-links in African swine fever virus DNA.

African swine fever virus DNA sediments in neutral sucrose density gradients as a single component with a sedimentation coefficient of 60S. In alkaline sucrose density gradients, this material shows two components with sedimentation coefficients of 85S and 95S, respectively. The sedimentation rate value of alkali-denatured virus DNA in neutral sucrose density gradients and the renaturation velocity of denatured DNA show that is reassociated much faster than expected from its genetic complexity. This behavior is compatible with the existence of interstrand cross-links in the molecule. We also present results which suggest that there are only a few such cross-links per molecule, that they are sensitive to S1 nuclease digestion, and that they are probably located next to the ends of the DNA.

African Swine Fever Virus

African swine fever: an epizootiological review with special reference to the South African situation.

The most important characteristics and the distribution of the viruses of African swine fever and hog cholera are reviewed. Both viruses were probably present simultaneously in South Africa in the first two decades of the century. While hog cholera was eradicated by 1918, African swine fever persists to the present day because it has a different epizootiology. The role played by wild pigs and the argasid tick (Ornithodoros moubata porcinus) in the epizootiology of African swine fever is discussed and an account of the outbreaks of the disease in South Africa from 1926 to 1974 is given. It appears that the disease in the Transvaal has had a cyclic occurrence.

Animals

Transcriptomic and Metabolomic Profiling Identifies a Core Gene-Metabolite Axis Driving African Swine Fever Virus Replication in the Soft Tick Ornithodoros lahorensis.

African swine fever virus (ASFV) causes an incurable swine disease with nearly 100% mortality, posing a catastrophic threat to global pig production. The soft tick Ornithodoros lahorensis acts as a critical biological vector that sustains persistent ASFV replication and mediates long-distance viral transmission, yet the molecular mechanisms governing ASFV-tick interplay remain poorly understood. Here, we integrated transcriptomics and metabolomics to systematically dissect molecular changes in O.&#xa0;lahorensis across three infection stages: Uninfected control, early infection (7&#x2009;days post-infection, dpi), and late persistent infection (21 dpi). Multi-omics integration revealed that ASFV extensively remodels tick host metabolism, predominantly activating purine/pyrimidine metabolism, lipid biosynthesis, and energy metabolism. We further characterized a conserved regulatory module consisting of 12 core genes and 8 signature metabolites that collectively support ASFV genome replication and virion assembly. Three hub metabolic genes (TK1, ATP5F1B, and IMPDH) were selected for functional validation via siRNA silencing in ticks; individual gene silencing suppressed ASFV loads by 89.2%, 91.5%, and 87.8%, respectively (p&#x2009;<&#x2009;0.001***). This work represents the first comprehensive multi-omics investigation of ASFV infection in O. lahorensis. We identified tick-specific molecular targets to block vector-mediated ASFV spread and established a standardized multi-omics analytical pipeline for tick-virus interaction research. Our findings elucidate the mechanistic basis of long-term ASFV persistence in soft ticks and deliver novel actionable clues for developing vector-targeted ASF intervention strategies.

Animals

The association of African swine fever virus with blood components of infected pigs.

The distribution of African swine fever virus (ASFV) in whole blood, plasma, red blood cells (RBC) and white blood cell (WbC) sub-populations was determined in pigs infected with virulent virus. Changes in the RBC and WBC populations were also examined. Total WBC counts decreased and RBC numbers remained unchanged during the course of the disease. The number of circulating lymphocytes decreased whilst neutrophil numbers increased owing to the replacement of mature forms by juveniles. Virus was present in all major blood fractions and was associated with equivalent numbers of both RBC and WBC. However, 90 per cent of the virus in whole blood was associated with RBC. Of the WBC subpopulations, virus was definitely associated with lymphocytes and possibly neutrophils.

African Swine Fever

African swine fever: pathogenicity and immunogenicity of two non-haemadsorbing viruses.

The virulence of 2 non-haemadsorbing African swine fever virus isolates were compared with 2 haemodsorbing viruses. While 3 of these isolates usually produced acute death in pigs, 1 non-haemadsorbing virus caused either a fatal infection with an extended course, or few or no obvious signs of infection. Pigs that survived infection with the latter virus were resistant to the lethal effects of the other 3 strains as well as to a pool of 7 isolates made from Ornithodorus porcinus porcinus (senus Walton, 1964) and warthog obtained in the Northern Transvaal.

African Swine Fever

Requirement of cell nucleus for African swine fever virus replication in Vero cells.

The role of the cell nucleus in the development of African swine fever virus in Vero cells has been studied. No viral growth could be detected in enucleated cells under conditions that allow normal development of Sindbis virus. Furthermore, African swine fever virus DNA synthesis was inhibited more than 95% after infection of enucleated Vero cells as compared with normal cells.

African Swine Fever Virus

Synthesis of DNA in cells infected with African swine fever virus.

Incorporation of 14C-thymidine by cells infected with African swine fever virus (ASFV) occurs in the nucleus. Part of this DNA is transferred to the cytoplasm and becomes resistant to DNAse. The nuclear fraction washed with Triton X100 retained all labeled DNA and was able to synthesize viral and cellular DNA under in vitro conditions in the presence of the four deoxynucleoside triphosphates, Mg+2, and sucrose. Under similar conditions nuclei from uninfected cells synthesized very little DNA.

African Swine Fever Virus

The growth of virulent African swine fever virus in pig monocytes and macrophages.

The replication of virulent African swine fever virus (ASFV) in cultures of monocytes and macrophages derived from pig bone marrow (PBM) and pig leukocyte (PL) cells was investigated by light microscopy, immunofluorescence, haemadsorption and infective virus release. Monocytes showed a high rate of infection and complete destruction within 2 to 3 days, whereas macrophages had only a very low level of infection and survived to form persistently infected cultures. These observations may explain the decrease in sensitivity of PBM and PL cells for ASFV assay after extended periods of incubation and suggest that the macrophage may be one of the cell types concerned with virus persistence in the pig.

African Swine Fever Virus

A solid-phase enzyme linked immunosorbent assay for the detection of African swine fever virus antigen and antibody.

A solid-phase enzyme-linked immunosorbent assay was developed to measure both African swine fever virus (ASFV) antigen and antibody. Experiments showed it to be reproducible and able to detect limiting antigen concentrations of 50--500 HAD50/ml. The assay was more sensitive than those used at present to detect ASFV antibody and it is suggested that it might be of great diagnostic use in countries where African swine fever has recently appeared.

African Swine Fever Virus

First detection of African swine fever in a swine farm in Taiwan.

INTRODUCTION: African swine fever (ASF) is a highly contagious, high-consequence transboundary animal disease that poses a critical threat to global swine production and agricultural economics. Since its emergence in China in 2018, ASF has spread to over 20 Asia-Pacific countries, causing significant economic disruption. While Taiwan previously detected ASF virus several times in dead pigs drifting offshore, or in illegal pork-related products brought by international tourists and inspected at airport and seaport border controls, no local domestic swine farm had tested positive before this October 2025 ASF event. However, maintaining this disease-free status requires constant vigilance against evolving regional biosecurity threats. METHODS: The first ASF detection in a domestic swine herd in Taichung City, Taiwan, reported on October 22, 2025, was triggered by abnormal alerts of the monitoring system in the rendering plant; the index farm captured a cumulative herd mortality rate of 35.2% that exceeded the predefined threshold (3% mortality daily in the nursery to finish pigs). Five finisher pigs were submitted for diagnostic evaluation, which subsequently confirmed ASFV infection via real-time PCR, pathological examination, immunohistochemistry, virus isolation, and whole-genome sequencing. RESULTS: Affected pigs showed clinical signs including wheezing, sudden death, nasal bleeding, uncoagulated blood in the nostrils, and mild hemorrhage on the skin surface of the neck, abdomen, and buttocks. Histopathological examination revealed severe multisystemic hemorrhagic lesions. Based on assay results of the P72, P54, P30, and CD2v genes, and whole-genome sequence, phylogenetic analysis confirmed that the isolate (ASFV/TWN/2025) is a genotype I/II recombinant strain, most similar to prevailing strains isolated in China and Vietnam, sharing 99.95%-99.97% and 99.92%-99.97% nucleotide similarity of whole genome sequence, respectively. DISCUSSION: Upon confirmation of the ASF case, authorities immediately implemented a nationwide swine movement standstill for 15 days to mitigate transmission risk. To date, no secondary cases have been detected. This article details the early monitoring and rapid diagnosis process of the first case of ASF infection in a farm in Taiwan, and highlights the information from this case to provide lessons for disease diagnosis and prevention in ASF-free areas.

African swine fever

The replication of virulent and attenuated strains of African swine fever virus in porcine macrophages.

The replication of virulent and attenuated strains of African swine fever virus (ASFV) was studied in pure cultures of swine macrophages. To ensure complete destruction of the macrophage monolayers about 50--100 times more virulent ASFV was needed than attenuated virus although both isolates could be used to establish persistently infected cultures. Interferon did not appear to influence virus yields from such cultures. Fluorescent and electron microscopy studies of infected macrophages suggested that the cycle of infection of the two isolates was different.

African Swine Fever Virus

Nanopore-based epigenomic profiling reveals the absence of widespread CpG methylation in the African swine fever virus genome.

DNA methylation is a critical epigenetic mechanism implicated in regulating replication and transcription in DNA viruses. However, the epigenetic landscape of African swine fever virus (ASFV), a large double-stranded DNA virus infecting pigs, remains controversial. Here, we systematically profiled the DNA methylome of the first ASFV strain isolated in Hong Kong (HK_NT_202103) using Oxford Nanopore Technologies (ONT) R10.4.1 sequencing. We employed a paired design: native whole-genome sequencing (WGS) against a methylation-free whole-genome amplification (WGA) control. Using conservative thresholds, we found no evidence of 5-methylcytosine (5mC), especially typical CpG methylation, across the viral genome. Importantly, clear CpG methylation signals were successfully detected in the host genome from WGS data, confirming the functionality of the workflow to detect 5mC at CG sites. While widespread 5mC seems absent, a small number of putative N6-methyladenine (6mA) loci were identified. A specific 6mA candidate exhibited raw ionic current disruptions and gene-level intersection with another ASFV isolate (CAS19-01/2019), although it lacked single-base consensus across different methylation callers or between the two isolates. Although our biological findings are restricted to a single isolate under specific experimental conditions, this study introduces a novel, highly rigorous ONT framework for viral epigenomics research. Furthermore, the absence of ASFV CpG methylation indicates that host CpG-depletion remains a viable strategy for viral metagenomic enrichment. Ultimately, our work offers a critical methodological baseline for ASFV surveillance and highlights the necessity of targeted experimental validation for rare viral modifications.

African Swine Fever Virus