Background The circular replication-associated protein (Rep)-encoding single-stranded (CRESS) DNA virus emergence in diverse host has been associated with severe disease. Porcine circovirus-like virus (Po-Circo-like [PCL] virus) is a CRESS DNA virus, the prevalence and pathogenicity of which are rarely studied. Methods We obtained two blood samples, four faecal samples, and two intestinal samples from a pig farm suffered from diarrheal disease in the delivery room in September 2020 and attempted to isolate and identify a causative pathogen. Subsequently, only PCL virus was positive, and qRT-PCR was designed to detect the loading titre of PCL virus. We then initiated a heightened surveillance program on the pathogenicity and epidemiology of PCL virus. Results Six PCL virus strains, with severe diarrhoea and haemorrhagic enteritis, have been found in six different pig farms in Guangdong province, China. A multiple sequence alignment of these PCL viruses and bovine circovirus-like virus/CH showed a similarity of 92.5-94.8% for the Rep protein, indicating these PCL viruses are highly homologous to Bo-Circo-like virus associated with calf diarrhoea. There were striking similarities between the PCL virus and bovine circovirus-like virus outbreaks in aetiological settings and Genomic sequence. We found that 11.2% (20/178) of diarrhoea samples and 13.3% (6/45) of pig farms were positive for PCL virus, suggesting that PCL virus may have spread widely in Pig farms. Moreover, this article underscores the risk of PCL virus spilling over and adapting to new species. Conclusions Porcine circovirus-like virus was found to be associated with porcine diarrheal disease in China.

(1) Background: Group A rotaviruses (RVAs) are the primary cause of severe intestinal diseases in piglets. Porcine rotaviruses (PoRVs) are widely prevalent in Chinese farms, resulting in significant economic losses to the livestock industry. However, isolation of PoRVs is challenging, and their pathogenicity in piglets is not well understood. (2) Methods: We conduct clinical testing on a farm in Jiangsu Province, China, and isolate strains of porcine rotavirus by continuously passaging on MA104 cells. Subsequently, the pathogenicity of the isolated strain in piglets was investigated.The piglets of the PoRV-infection group were orally inoculated with 1 mL of 1.0 × 10⁶ TCID50 PoRV, whereas those of the mock-infection group were fed with an equivalent amount of DMEM. (3) Results: Fecal samples were collected from 28 piglets, all of which tested positive for RVAs. A G5P[23] genotype porcine rotavirus strain was successfully isolated from one of the positive samples and named RVA/Pig/China/JS/2023/G5P[23](JS). The genomic constellation of this strain was G5-P[23]-I5-R1-C1-M1-A8-N1-T1-E1-H1. Sequence analysis revealed that the genes VP3, VP7, NSP2, and NSP4 of the JS strain were closely related to human RVAs, whereas the remaining gene segments were closely related to porcine RVAs, indicating a reassortment between porcine and human strains. Furthermore, infection of 15-day-old piglets with the JS strain resulted in a diarrheal rate of 100% (8 of 8) and mortality rate of 37.5% (3 of 8).(4) Conclusion: The isolated G5P [23] genotype rotavirus strain, which exhibited strong pathogenicity in piglets, may have resulted from recombination between porcine and human strains. It may serve as a potential candidate strain for developing vaccines, and its immunogenicity can be tested in future studies.

Porcine epidemic diarrhea virus (PEDV) and transmissible gastroenteritis virus (TGEV), two economically devastating swine enteric coronaviruses, persistently threaten global pork production via severe clinical morbidity. Vaccination is the most effective means to prevent morbidity and mortality caused by porcine enteric coronavirus infection. Hence, the development of an effective and safe vaccination strategy to protect against both PEDV and TGEV infections is urgently needed. Here, we designed and produced trimerized full-length PEDV and TGEV spike (S) proteins using an efficient mammalian expression vector system in HEK293F cells and reported that compared with the PEDV spike protein (2 ⁷ ), the TGEV spike protein induces significantly more potent neutralizing antibodies (2 ¹² ). Moreover, transmission electron microscopy (TEM) with negative staining revealed that the TGEV spike protein exhibited excellent homogeneity. We subsequently used the TGEV S protein trimer as the backbone to display different domains of the PEDV S protein and successfully constructed seven spike protein chimeras. We further screened two chimeric S proteins through expression and mouse experiments, which included TGEV S-PEDV(D0/NTD) and TGEV S-PEDV(D0/NTD/CTD), in which the S protein of TGEV was replaced by the PEDV S D0-NTD and D0-NTD-CTD, respectively. Evaluation of the immunogenicity and efficacy of the chimeric S proteins in piglets confirmed that piglets in the TGEV S-PEDV(D0/NTD)-immunized group generated high levels of neutralizing antibodies (2 ^13.6 against TGEV; 2 ^5.8 against PEDV) against both PEDV and TGEV. Our findings suggest that the TGEV S-PEDV(D0/NTD) is a promising candidate for combating both PEDV and TGEV infections. IMPORTANCE: The design strategy of multivalent and multitarget single antigens facilitates the development of vaccines targeting PEDV and TGEV. Optimizing the presentation of PEDV core antigen epitopes on the basis of the S protein trimer structure will provide novel insights into the development of subunit vaccines. Here, we compared the immunogenicities of the TGEV S protein and PEDV S protein and then designed a bivalent chimeric S protein candidate, TGEV S-PEDV(D0/NTD), in which the corresponding segments on the TGEV S protein were replaced with D0-NTD domains from the PEDV S protein. We demonstrated that the TGEV S protein exhibits greater immunogenicity than the PEDV S protein and that TGEV S-PEDV(D0/NTD) induces broad-spectrum neutralization protection against PEDV and TGEV. Our results demonstrate the effectiveness of the chimeric S protein and provide a feasible method for the development of efficient bivalent subunit vaccines against PEDV and TGEV.

Porcine epidemic diarrhea virus (PEDV) is an enteric coronavirus that infects pigs of all ages. Notably, neonatal piglets (≤7 days old) in naïve herds can experience case fatality rates approaching 100%, with hallmark clinical signs of vomiting, diarrhea, and dehydration, thereby representing a major threat to swine herd health. Serum PEDV‐specific IgG serves as a key serological marker of vaccine‐induced herd immunity. Accordingly, we developed an indirect ELISA (iELISA) using Chinese hamster ovary (CHO) cell‐expressed recombinant PEDV spike (S) protein as the coating antigen. Checkerboard titration identified the optimal antigen‐coating concentration and serum dilution as 1 μg/mL and 1:300, respectively. Based on sample‐to‐positive (S/P) ratios from 50 PEDV‐negative sera, the diagnostic cutoff was set at 0.796, corresponding to the mean plus 3 standard deviation (SD) of the negative control group ( n = 50). The intra‐ and interassay coefficients of variation (CVs) were both <6.30%, indicating good precision (repeatability) and reproducibility. Specificity testing showed no detectable cross‐reactivity with sera positive for five common porcine viruses (porcine deltacoronavirus (PDCoV), transmissible gastroenteritis virus (TGEV), porcine group A rotavirus (PoRVA), porcine reproductive and respiratory syndrome virus (PRRSV), and African swine fever virus (ASFV)). Moreover, PEDV‐specific IgG measured by iELISA correlated strongly with virus‐neutralizing (VN) antibody titers ( r = 0.95), and both metrics were associated with protective outcomes at the herd level. Collectively, this assay provides an effective tool for evaluating protective immunity following PEDV vaccination.

Porcine epidemic diarrhea virus (PEDV) remains a major threat to the global swine industry, with genotype II (GII) strains causing nearly 100% mortality in neonatal piglets. Current vaccination strategies and biosafety control measures provide limited cross-protection and may pose potential biosafety risks, highlighting the urgent need for safer and more effective preventive approaches. In this study, we evaluated a spike (S) protein subunit vaccine administered via needle-free intradermal (ID) immunization and assessed its capacity to induce maternally derived passive protection. ID immunization elicited robust PEDV-specific IgG and neutralizing antibody responses comparable to those induced by conventional intramuscular (IM) injection, with immune responses maintained for more than four months. High levels of IgG and neutralizing antibodies were detected in the serum and colostrum of vaccinated sows, and piglets born to these sows exhibited only mild or no diarrhea following virulent PEDV challenge. Viral shedding and intestinal viral loads were markedly reduced, and intestinal lesions were nearly completely prevented. Overall, this needle-free ID vaccination strategy conferred more than 95% passive protection in neonatal piglets while improving animal welfare and vaccination efficiency. These findings demonstrate that ID delivery of an S protein subunit vaccine represents a promising and practical alternative to conventional IM vaccination for effective PEDV control.