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Journal of Nanobiotechnology May 2024Widespread distribution of porcine epidemic diarrhea virus (PEDV) has led to catastrophic losses to the global pig farming industry. As a result, there is an urgent need...
Widespread distribution of porcine epidemic diarrhea virus (PEDV) has led to catastrophic losses to the global pig farming industry. As a result, there is an urgent need for rapid, sensitive and accurate tests for PEDV to enable timely and effective interventions. In the present study, we develop and validate a floating gate carbon nanotubes field-effect transistor (FG CNT-FET)-based portable immunosensor for rapid identification of PEDV in a sensitive and accurate manner. To improve the affinity, a unique PEDV spike protein-specific monoclonal antibody is prepared by purification, and subsequently modified on FG CNT-FET sensor to recognize PEDV. The developed FET biosensor enables highly sensitive detection (LoD: 8.1 fg/mL and 10 TCID/mL for recombinant spike proteins and PEDV, respectively), as well as satisfactory specificity. Notably, an integrated portable platform consisting of a pluggable FG CNT-FET chip and a portable device can discriminate PEDV positive from negative samples and even identify PEDV and porcine deltacoronavirus within 1 min with 100% accuracy. The portable sensing platform offers the capability to quickly, sensitively and accurately identify PEDV, which further points to a possibility of point of care (POC) applications of large-scale surveillance in pig breeding facilities.
Topics: Porcine epidemic diarrhea virus; Animals; Swine; Biosensing Techniques; Nanotubes, Carbon; Limit of Detection; Immunoassay; Antibodies, Monoclonal; Transistors, Electronic; Swine Diseases; Spike Glycoprotein, Coronavirus; Coronavirus Infections; Antibodies, Viral; Equipment Design
PubMed: 38735951
DOI: 10.1186/s12951-024-02440-5 -
Infection, Genetics and Evolution :... Aug 2023Recent reports on identification of canine coronavirus (CCoV) in humans have emphasized the urgency to strengthen surveillance of animal CoVs. The fact that...
Recent reports on identification of canine coronavirus (CCoV) in humans have emphasized the urgency to strengthen surveillance of animal CoVs. The fact that recombinations between CCoV with feline, porcine CoVs brought about new types of CoVs indicated that more attention should be paid to domestic animals like dogs, cats and pigs, and the CoVs they carried. However, there are about ten kinds of CoVs that infect above animals, and thus representative CoVs with zoonotic potentials were considered in this study. Multiplex RT-PCR against CCoV, Feline coronavirus (FCoV), porcine deltacoronavirus and porcine acute diarrhea syndrome coronavirus was developed to investigate the prevalence of CoVs from domestic dogs in Chengdu, Southwest China. Samples from a total of 117 dogs were collected from a veterinary hospital, and only CCoV (34.2%, 40/117) was detected. Therefore, this study focused on CCoV and its characteristics of S, E, M, N and ORF3abc genes. Compared with CoVs that are capable of infecting humans, CCoV strains showed highest nucleotide identity with the novel canine-feline recombinant detected from humans (CCoV-Hupn-2018). Phylogenetic analysis based on S gene, CCoV strains were not only clustered with CCoV-II strains, but also closely related to FCoV-II strains ZJU1617 and SMU-CD59/2018. As for assembled ORF3abc, E, M, N sequences, CCoV strains had the closest relationship with CCoV-II (B203_GZ_2019, B135_JS_2018 and JS2103). What's more, specific amino acid variations were found, especially in S and N proteins, and some mutations were consistent with FCoV, TGEV strains. Altogether, this study provided a novel insight into the identification, diversification and evolution of CoVs from domestic dogs. It is of top priority to recognize zoonotic potential of CoVs, and continued comprehensive surveillance will help better understand the emergence, spreading, and ecology of animal CoVs.
Topics: Animals; Dogs; Cats; Humans; Swine; Coronavirus, Canine; Coronavirus Infections; Reverse Transcriptase Polymerase Chain Reaction; Phylogeny; Molecular Epidemiology; Mutation; Animals, Domestic; China; Dog Diseases
PubMed: 37295484
DOI: 10.1016/j.meegid.2023.105463 -
Microorganisms Feb 2024Porcine deltacoronavirus (PDCoV) has shown large-scale global spread since its discovery in Hong Kong in 2012. In this study, a total of 4897 diarrheal fecal samples...
Porcine deltacoronavirus (PDCoV) has shown large-scale global spread since its discovery in Hong Kong in 2012. In this study, a total of 4897 diarrheal fecal samples were collected from the Guangxi province of China from 2020 to 2023 and tested using RT-qPCR. In total, 362 (362/4897, 7.39%) of samples were positive for PDCoV. The S, M, and N gene sequences were obtained from 34 positive samples after amplification and sequencing. These PDCoV gene sequences, together with other PDCoV S gene reference sequences from China and other countries, were analyzed. Phylogenetic analysis revealed that the Chinese PDCoV strains have diverged in recent years. Bayesian analysis revealed that the new China 1.3 lineage began to diverge in 2012. Comparing the amino acids of the China 1.3 lineage with those of other lineages, the China 1.3 lineage showed variations of mutations, deletions, and insertions, and some variations demonstrated the same as or similar to those of the China 1.2 lineage. In addition, recombination analysis revealed interlineage recombination in CHGX-MT505459-2019 and CHGX-MT505449-2017 strains from Guangxi province. In summary, the results provide new information on the prevalence and evolution of PDCoV in Guangxi province in southern China, which will facilitate better comprehension and prevention of PDCoV.
PubMed: 38399820
DOI: 10.3390/microorganisms12020416 -
BMC Veterinary Research Sep 2023Porcine epidemic diarrhea virus (PEDV) and porcine delta-coronavirus (PDCoV) are economically important pathogens that cause diarrhea in sows and acute death of newborn...
BACKGROUND
Porcine epidemic diarrhea virus (PEDV) and porcine delta-coronavirus (PDCoV) are economically important pathogens that cause diarrhea in sows and acute death of newborn piglets. Moreover, the emerging PDCoV was reported to infect children. The current situation is that vaccine prevention has not met expectations, and emergency containment strategies following outbreaks cannot prevent the damages and losses already incurred. Therefore, a more sensitive detection method, that is both convenient and enables accurate and effective sequencing, that will provide early warning of PEDV and PDCoV is necessary. This will enable active, effective, and comprehensive prevention and control, which will possibly reduce disease occurrences.
RESULTS
Duplex nested RT-PCR (dnRT-PCR) is an ideal method to achieve early warning and monitoring of PEDV and PDCoV diseases, and to additionally investigate any molecular epidemiological characteristics. In this study, two pairs of primers were designed for each virus based upon the highly conserved N protein sequences of both PEDV and PDCoV strains retrieved from the NCBI Genbank. After optimization of the reaction conditions, the dnRT-PCR assay amplified a 749-bp fragment specific to PEDV and a 344-bp fragment specific to PDCoV. Meanwhile, the specificity and sensitivity of the primers and clinical samples were tested to verify and establish this dnRT-PCR method. The limit of detection (LoD)for both PEDV and PDCoV was 10 copies/µL. The results showed that among 251 samples, 1 sample contained PEDV infection, 19 samples contained a PDCoV infection, and 8 samples were infected with both viruses, following the use of dnRT-PCR. Subsequently, the positive samples were sent for sequencing, and the sequencing results confirmed that they were all positive for the viruses detected using dnRT-PCR, and conventional RT-PCR detection was conducted again after the onset of disease. As these results were consistent with previous results, a detection method for PEDV and PDCoV using dnRT-PCR was successfully established. In conclusion, the dnRT-PCR method established in this study was able to detect both PEDV and PDCoV, concomitantly.
CONCLUSIONS
The duplex nested RT-PCR method represents a convenient, reliable, specific, sensitive and anti-interference technique for detecting PEDV and PDCoV, and can additionally be used to simultaneously determine the molecular epidemiological background.
Topics: Animals; Swine; Female; Coronavirus; Porcine epidemic diarrhea virus; Reverse Transcriptase Polymerase Chain Reaction; Coronavirus Infections; Polymerase Chain Reaction; DNA Primers
PubMed: 37684673
DOI: 10.1186/s12917-023-03708-y -
Porcine Health Management Mar 2024Diarrheal diseases caused by viral agents have led to a great morbidity, mortality, and economic loss in global pig industry. Porcine epidemic diarrhea virus (PEDV),...
BACKGROUND
Diarrheal diseases caused by viral agents have led to a great morbidity, mortality, and economic loss in global pig industry. Porcine epidemic diarrhea virus (PEDV), transmissible gastroenteritis virus (TGEV), porcine deltacoronavirus (PDCoV), and group A porcine rotavirus (RVA) are main causative agents of swine viral diarrhea with similar clinical signs on Chinese farms and their co-infection is also common. However, it is still lack of a convenient method to detect these four agents.
METHODS
A TaqMan multiplex qPCR method was developed to detect PEDV, TGEV, PDCoV, and RVA, simultaneously. This method was then applied to investigate 7,342 swine fecal samples or rectal swabs, as well as 1,246 swine intestinal samples collected from 2075 farms in China in 2022.
RESULTS
Minimum detection limits of this method were 3 copies/µL for PEDV, 4 copies/µL for TGEV, 8 copies/µL for RVA, and 8 copies/µL for PDCoV, suggesting a good sensitivity. No signals were observed by using this method detecting other viral agents commonly prevalent in pigs, which is suggestive of a good specificity. Application of this method on investigating clinical samples demonstrated a relatively high positive rate for PEDV (22.21%, 1907/8588) and RVA (44.00%, 3779/8588). In addition, co-infection between PEDV and RVA was observed on 360 investigated farms, accounting for 17.35% (360/2075) of the farms where co-infection events were screened.
CONCLUSIONS
A TaqMan multiplex qPCR method targeting PEDV, TGEV, PDCoV, and RVA was developed in this study. This method demonstrated a good specificity and sensitivity on investigating these four common viruses responsible for viral diarrhea on Chinese pig farms, which represents a convenient method for the monitoring and differential diagnosis of swine viral diarrhea.
PubMed: 38444040
DOI: 10.1186/s40813-024-00364-y -
Veterinary Research Jun 2024Porcine deltacoronavirus (PDCoV) is an enteropathogenic coronavirus that has been reported to use various strategies to counter the host antiviral innate immune...
Porcine deltacoronavirus (PDCoV) is an enteropathogenic coronavirus that has been reported to use various strategies to counter the host antiviral innate immune response. The cGAS-STING signalling pathway plays an important role in antiviral innate immunity. However, it remains unclear whether PDCoV achieves immune evasion by regulating the cGAS-STING pathway. Here, we demonstrated that the nonstructural protein 2 (nsp2) encoded by PDCoV inhibits cGAS-STING-mediated type I and III interferon (IFN) responses via the regulation of porcine STING (pSTING) stability. Mechanistically, ectopically expressed PDCoV nsp2 was found to interact with the N-terminal region of pSTING. Consequently, pSTING was degraded through K48-linked ubiquitination and the proteasomal pathway, leading to the disruption of cGAS-STING signalling. Furthermore, K150 and K236 of pSTING were identified as crucial residues for nsp2-mediated ubiquitination and degradation. In summary, our findings provide a basis for elucidating the immune evasion mechanism of PDCoV and will contribute to the development of targets for anti-coronavirus drugs.
Topics: Animals; Swine; Viral Nonstructural Proteins; Deltacoronavirus; Swine Diseases; Membrane Proteins; Coronavirus Infections; Interferon Type I; Immunity, Innate; HEK293 Cells; Immune Evasion; Ubiquitination
PubMed: 38886840
DOI: 10.1186/s13567-024-01330-w -
Viruses Jul 2023Swine coronaviruses (CoVs) have been found to cause infection in humans, suggesting that Suiformes might be potential intermediate hosts in CoV transmission from their...
Swine coronaviruses (CoVs) have been found to cause infection in humans, suggesting that Suiformes might be potential intermediate hosts in CoV transmission from their natural hosts to humans. The present study aims to establish convolutional neural network (CNN) models to predict host adaptation of swine CoVs. Decomposing of each and sequence was performed with dinucleotide composition representation (DCR) and other traits. The relationship between CoVs from different adaptive hosts was analyzed by unsupervised learning, and CNN models based on DCR of and were built to predict the host adaptation of swine CoVs. The rationality of the models was verified with phylogenetic analysis. Unsupervised learning showed that there is a multiple host adaptation of different swine CoVs. According to the adaptation prediction of CNN models, swine acute diarrhea syndrome CoV (SADS-CoV) and porcine epidemic diarrhea virus (PEDV) are adapted to Chiroptera, swine transmissible gastroenteritis virus (TGEV) is adapted to Carnivora, porcine hemagglutinating encephalomyelitis (PHEV) might be adapted to Primate, Rodent, and Lagomorpha, and porcine deltacoronavirus (PDCoV) might be adapted to Chiroptera, Artiodactyla, and Carnivora. In summary, the DCR trait has been confirmed to be representative for the CoV genome, and the DCR-based deep learning model works well to assess the adaptation of swine CoVs to other mammals. Suiformes might be intermediate hosts for human CoVs and other mammalian CoVs. The present study provides a novel approach to assess the risk of adaptation and transmission to humans and other mammals of swine CoVs.
Topics: Swine; Animals; Humans; Coronavirus; Chiroptera; Phylogeny; Deep Learning; Swine Diseases; Coronavirus Infections; Porcine epidemic diarrhea virus; Risk Assessment; Carnivora
PubMed: 37515242
DOI: 10.3390/v15071556 -
Frontiers in Veterinary Science 2024Porcine deltacoronavirus (PDCoV), an emerging swine enteropathogenic coronavirus with worldwide distribution, mainly infects newborn piglets with severe diarrhea,...
INTRODUCTION
Porcine deltacoronavirus (PDCoV), an emerging swine enteropathogenic coronavirus with worldwide distribution, mainly infects newborn piglets with severe diarrhea, vomiting, dehydration, and even death, causing huge economic losses to the pig industry. However, the underlying pathogenic mechanisms of PDCoV infection and the effects of PDCoV infection on host transcripts and metabolites remain incompletely understood.
METHODS
This study investigated a combined transcriptomic and metabolomic analysis of porcine intestinal epithelial cells (IPEC-J2) following PDCoV infection by LC/MS and RNA-seq techniques. A total of 1,401 differentially expressed genes and 254 differentially accumulated metabolites were detected in the comparison group of PDCoV-infected vs. mock-infected.
RESULTS AND DISCUSSION
We found that PDCoV infection regulates gene sets associated with multiple signaling pathways, including the neuroactive ligand-receptor interaction, cytokine-cytokine receptor interaction, MAPK signaling pathway, chemokine signaling pathway, ras signaling pathway and so on. Besides, the metabolomic results showed that biosynthesis of cofactors, nucleotide metabolism, protein digestion and absorption, and biosynthesis of amino acid were involved in PDCoV infection. Moreover, integrated transcriptomics and metabolomics analyses revealed the involvement of ferroptosis in PDCoV infection, and exogenous addition of the ferroptosis activator erastin significantly inhibited PDCoV replication. Overall, these unique transcriptional and metabolic reprogramming features may provide a better understanding of PDCoV-infected IPEC-J2 cells and potential targets for antiviral treatment.
PubMed: 38855411
DOI: 10.3389/fvets.2024.1359547 -
Frontiers in Veterinary Science 2024-glycosylation is a highly conserved glycan modification that plays crucial roles in various physiological processes, including protein folding, trafficking, and signal...
-glycosylation is a highly conserved glycan modification that plays crucial roles in various physiological processes, including protein folding, trafficking, and signal transduction. Porcine deltacoronavirus (PDCoV) poses a newly emerging threat to the global porcine industry. The spike protein of PDCoV exhibits a high level of -glycosylation; however, its role in viral infection remains poorly understood. In this study, we applied a lentivirus-based entry reporter system to investigate the role of -glycosylation on the viral spike protein during PDCoV entry stage. Our findings demonstrate that -glycosylation at positions 652 and 661 of the viral spike protein significantly reduces the infectivity of PDCoV pseudotyped virus. Overall, our results unveil a novel function of -glycosylation in PDCoV infection, highlighting its potential for facilitating the development of antiviral strategies.
PubMed: 38872801
DOI: 10.3389/fvets.2024.1430113 -
Microorganisms Apr 2024This study was conducted to elucidate the intestinal damage induced by the IPEC-J2 cell culture-passaged PDCoV. The results showed that PDCoV disrupted the intestinal...
This study was conducted to elucidate the intestinal damage induced by the IPEC-J2 cell culture-passaged PDCoV. The results showed that PDCoV disrupted the intestinal structure and increased intestinal permeability, causing abnormalities in mucosal pathology. Additionally, PDCoV induced an imbalance in the intestinal flora and disturbed its stability. Microbial community profiling revealed bacterial enrichment (e.g., Proteobacteria) and reduction (e.g., Firmicutes and Bacteroidetes) in the PDCoV-inoculated piglet model. In addition, metabolomics analysis indicated that 82 named differential metabolites were successfully quantified, including 37 up-regulated and 45 down-regulated metabolites. Chenodeoxycholic acid, sphingosine, and oleanolic aldehyde levels were reduced in PDCoV-inoculated piglets, while phenylacetylglycine and geranylgeranyl-PP levels were elevated. Correlation analysis indicated a negative correlation between and choline, succinic acid, creatine, phenyllactate, and hippuric acid. Meanwhile, was positively correlated with acetylcholine, L-Glutamicacid, and N-Acetylmuramate. , , , and were negatively and positively correlated with sphingosine, respectively. These data suggested PDCoV-inoculated piglets exhibited significant taxonomic perturbations in the gut microbiome, which may result in a significantly altered metabolomic profile.
PubMed: 38792704
DOI: 10.3390/microorganisms12050874