Porcine transmissible gastroenteritis virus (TGEV) is a highly contagious pathogen causing severe diarrhea in pigs, particularly piglets, leading to significant economic losses. Distinguishing TGEV from the genetically similar porcine respiratory coronavirus (PRCV) remains challenging due to their high genomic homology. In this study, we developed a one–pot assay combining recombinase-aided amplification (RAA) and CRISPR/Cas13a technology, targeting the TGEV S gene. This method was optimized for sensitivity and specificity, with orthogonal tests determining the optimal reagent concentrations. The assay achieved a detection limit of 4.13 copies/µL within 40 min at 37 °C, demonstrating no cross-reactivity with other porcine viruses. Clinical validation on 140 samples showed 100% concordance with RT–qPCR and RT–PCR results. Since the established method is completed in a single reaction tube, it eliminates the need for step-by-step operations, simplifying the process and reducing the risk of cross–contamination and false positives in subsequent tests. Overall, this assay shows promising potential for TGEV detection.

Porcine epidemic diarrhea virus (PEDV) continues to undergo persistent genetic evolution and remains highly prevalent in pig herds worldwide. The resulting porcine epidemic diarrhea (PED) causes substantial economic losses to the global commercial swine industry. In this study, 296 clinical specimens were collected from diarrheic piglets across nine districts and counties of Chongqing between 2022–2024 and were screened using RT-qPCR. PEDV-positive specimens were detected in all sampling regions, with an overall positive rate of 48.31% (143/296), indicating widespread circulation and endemic persistence of PEDV throughout Chongqing. Phylogenetic analysis of the S gene from 15 representative field isolates revealed that all strains clustered within the G2c sublineage, implying that G2c has become the predominant epidemic lineage circulating in this region in recent years. Sequence analysis showed that the nucleotide identities of the S gene ranged from 93.37%–94.09% compared with the classic CV777 strain and from 96.79%–97.67% compared with the variant AJ1102 strain. Corresponding amino acid similarities were 92.88%–93.56% and 97.25%–98.05%, respectively. Notably, clear recombination footprints were identified in 11 sequenced isolates, all exhibiting highly identical recombination patterns, likely resulting from crossover events between the major parental strain FR/001/2014 and the minor parental strain CH/GDGZ/2012. Furthermore, ten distinct amino acid substitutions including F536L, D566E and L573S were identified within the COE antigenic domain of the S protein, whereas the SS2, SS6 and 2C10 domains remained highly conserved. Four isolates possessed four novel potential N-glycosylation motifs, including 300 NKTI and 787 NFSV, while two isolates concurrently lost the conserved 341 NLSF glycosylation site. These genetic alterations may influence viral pathogenicity and immune evasion capacity, thereby potentially contributing to the continued predominance and spread of the G2c sublineage in China. Together, this study expands current epidemiological knowledge of PEDV in inland Southwest China, and confirms G2c strains as the dominant circulating variants in local pig populations. These findings provide important molecular insights into the epidemiological trends and evolutionary dynamics of PEDV and offer a valuable reference for the development of more effective prevention and control strategies.

Rapid and accurate pathogen detection is crucial to mitigating the significant economic impacts of infectious animal diseases. Recombinase-aided amplification (RAA) has emerged as a highly promising isothermal diagnostic tool, characterized by its rapid turnaround time, robust amplification efficiency, and single-copy analytical sensitivity. In veterinary diagnostics, RAA has evolved from single-pathogen proof-of-concept assays to become a promising technology for point-of-care scenarios. This review critically evaluates the integration of RAA with real-time fluorescence, CRISPR-Cas systems, lateral flow dipsticks (LFDs), and suspension microarray technologies, highlighting their impact on assay sensitivity and point-of-care (POC) applicability. Despite recent advancements, including encouraging progress in multiplexing and expanded applications in companion animal medicine, inherent challenges such as matrix effects in crude clinical samples, aerosol contamination risks, and commercialization barriers remain. By benchmarking RAA against traditional molecular methods and structurally similar technologies like recombinase polymerase amplification (RPA), this review outlines future trajectories for optimizing RAA platforms, aiming to maximize their utility in routine veterinary surveillance and epidemiological control.