Porcine epidemic diarrhea virus (PEDV) causes devastating enteric disease that inflicts huge economic damage on the swine industry worldwide. A safe and highly effective PEDV vaccine that contains only the virus-neutralizing epitopes (not enhancing epitope), as well as a ready-to-use PEDV neutralizing antibody for the passive immunization of PEDV vulnerable piglets (during the first week of life) are needed, particularly for PEDV-endemic farms. In this study, we generated monoclonal antibodies (mAbs) to the recombinant S1 domain of PEDV spike (S) protein and tested their PEDV neutralizing activity by CPE-reduction assay. The mAb secreted by one hybrodoma clone (A3), that also bound to the native S1 counterpart from PEDV-infected cells (tested by combined co-immunoprecipitation and Western blotting), neutralized PEDV infectivity. Epitope of the neutralizing mAb (mAbA3) locates in the S1A subdomain of the spike protein, as identified by phage mimotope search and multiple sequence alignment, and peptide binding-ELISA. The newly identified epitope is shared by PEDV G1 and G2 strains and other alphacoronaviruses. In summary, mAbA3 may be useful as a ready-to-use antibody for passive immunization of PEDV-susceptible piglets, while the novel neutralizing epitope, together with other, previously known protective epitopes, have potential as an immunogenic cocktail for a safe, next-generation PEDV vaccine.

Background and Aim: Acinetobacter baumannii is a multidrug-resistant (MDR) pathogen notorious for its biofilm formation and persistence in clinical and veterinary settings. Its resistance is exacerbated by quorum sensing (QS) pathways that regulate virulence and biofilm maturation. Disrupting QS and biofilm integrity using plant-derived compounds presents a promising alternative to traditional antibiotics. This study aimed to evaluate the antibiofilm and anti-QS potential of Paederia foetida Linn. ethanolic extract against A. baumannii, integrating gas chromatography–mass spectrometry (GC-MS) profiling, molecular docking, and in vitro assays. Materials and Methods: Leaves of P. foetida were extracted with ethanol and analyzed by GC-MS to identify major bioactive constituents. Molecular docking was conducted against five QS and biofilm-associated A. baumannii proteins (AF-A0A7S8WE28-F1-v4, AF-A0A059ZL64-F1-v4, AF-Q2VSW6-F1-v4, AF-A0A2P1B9S4-F1-v4, and AF-A0A5P9VY74-F1-v4). Absorption, distribution, metabolism, excretion, and toxicity (ADMET) profiles and drug-likeness of key compounds were assessed in silico. Antimicrobial activity was determined by broth microdilution (minimum inhibitory concentration [MIC]/minimum bactericidal concentration [MBC]), and biofilm inhibition was evaluated through crystal violet microtiter assays. Morphological damage was examined using field emission scanning electron microscopy (FE-SEM). Results: GC-MS identified 30 phytoconstituents, with 5-hydroxymethyl-2-furaldehyde, 4H-pyran-4-one derivative, and eugenol as predominant compounds. Eugenol exhibited the highest binding affinity, particularly with AbaR (−6.3 kcal/mol). The extract showed significant antimicrobial activity (MIC = 7.81 mg/mL; MBC = 31.25 mg/mL) and dose-dependent inhibition of biofilm biomass (p < 0.001). FE-SEM imaging confirmed dose-responsive membrane damage and disruption of the biofilm. ADMET predictions revealed favorable oral bioavailability and low toxicity for selected compounds. Conclusion: P. foetida extract exhibits potent antibacterial, anti-QS, and antibiofilm activity against MDR A. baumannii, supported by its phytochemical diversity, favorable pharmacokinetics, and strong protein-ligand interactions. These findings suggest its promise as a plant-derived therapeutic aligned with the One Health framework to combat antimicrobial resistance in both human and veterinary medicine. Keywords: Acinetobacter baumannii, absorption, distribution, metabolism, excretion, and toxicity, biofilm disruption, eugenol, gas chromatography–mass spectrometry, molecular docking, One Health, Paederia foetida, quorum sensing inhibition.

Porcine epidemic diarrhea virus (PEDV) infection is an important acute diarrheal disease of swine that results in economic and industrial losses worldwide. The clinical manifestations in infected piglets are severe diarrhea, dehydration with milk curd indigestion, leading to death. The diagnosis of PEDV is essential for monitoring and managing the disease. PEDV can be detected and identified by serology and the nucleic acid of the virus in clinical samples. Therefore, a novel isothermal amplification and detection technique, reverse transcription-recombinase polymerase amplification couple nucleic acid lateral flow (RT-RPA-NALF) was developed for the rapid detection of PEDV. Qualitative reverse transcription-polymerase chain reaction (RT-qPCR) was established as the gold standard assay to compare results. Specific primer pairs and probes were designed, and RT-RPA conditions were optimized to amplify the M gene of PEDV. The established RT-RPA-NALF assay could finish in 25 min at a temperature of 42 °C and the amplicon interpreted by visual detection. The developed RT-RPA-NALF assay was specific to the M gene of PEDV, did not detect other common swine diarrhea pathogens, and showed minimal detection at 10² TCID₅₀/mL PEDV. The RT-RPA-NALF assay can detect PEDV in 5 simulated fecal samples. Furthermore, in 60 clinical fecal samples, the results of RT-RPA-NALF correlated with RT-qPCR assay, which provides sensitivity of 95.65% and specificity of 100%, with a coincident rate of 98.33%. The rapid RT-RPA-NALF is simple and rapid, increases high sensitivity, and can be used in the field.

During the COVID-19 pandemic, the parasitology laboratories dealing with fecal samples for the diagnosis of gastrointestinal parasitic infections are confronting the unsaved virus-containing samples. To allow for safe downstream processing of the fecal samples, a protocol for preparing a fecal smear is urgently needed. Formalin was tested with or without isotonic forms for virus inactivation using porcine epidemic diarrhea virus (PEDV) as a representative, as it belongs to the Coronaviridae family. The results revealed complete inactivation activity of 10% formalin and 10% isotonic formalin on coronavirus after 5 min of treatment at room temperature. Both also inhibited Naegleria fowleri growth after 5 min of treatment at 37 °C without disruption of the structure. In addition to these key findings, it was also found that isotonic formalin could stabilize both red and white blood cells when used as a solution to prepare fecal smears comparable to the standard method, highlighting its value for use instead of 0.9% normal saline solution for the quantification of blood cells without active virus. The 10% isotonic formalin is useful to safely prepare a fecal smear for the diagnosis of parasites and other infections of the gastrointestinal tract during the COVID-19 pandemic.