Phylogeography and evolutionary dynamics analysis of porcine delta-coronavirus with host expansion to humans.

From 2003 onwards, three pandemics have been caused by coronaviruses: severe acute respiratory syndrome coronavirus (SARS-CoV); middle east respiratory syndrome coronavirus (MERS-CoV); and, most recently, SARS-CoV-2. Notably, all three were transmitted from animals to humans. This would suggest that animals are potential sources of epidemics for humans. The emerging porcine delta-coronavirus was reported to infect children. This is a red flag that marks the ability of PDCoV to break barriers of cross-species transmission to humans. Therefore, we conducted molecular genetic analysis of global clade PDCoV to characterize spatiotemporal patterns of viral diffusion and genetic diversity. PDCoV was classified into three major lineages, according to distribution and phylogenetic analysis of PDCoV. It can be inferred based on the analysis results of the currently known PDCoV strains that PDCoV might originate in Asia. We also selected six special spike amino acid sequences to align and analyze to find seven significant mutation sites. The accumulation of these mutations may enhance dynamic movements, accelerating spike protein membrane fusion events and transmission. Altogether, our study offers a novel insight into the diversification, evolution, and interspecies transmission and origin of PDCoV and emphasizes the need to study the zoonotic potential of the PDCoV and comprehensive surveillance and enhanced biosecurity precautions for PDCoV.

Buffalo-Origin Seneca Valley Virus in China: First Report, Isolation, Genome Characterization, and Evolution Analysis.

Pigs are the main host of Seneca Valley virus (SVV), previously known as Senecavirus A (SVA). Pigs affected by SVV have vesicles in the nose, hooves, and limp and may cause death in some severe cases. Occasionally, SVV has also been detected in mice, houseflies, environmental equipment, and corridors in pig farms. Moreover, it was successfully isolated from mouse tissue samples. In this study, an SVV strain (SVA/GD/China/2018) was isolated from a buffalo with mouth ulcers in the Guangdong province of China using seven mammalian cell lines (including BHK-21, NA, PK-15, ST, Vero, Marc-145, and MDBK). The genome of SVA/GD/China/2018 consists of 7,276 nucleotides. Multiple-sequence alignment showed that SVA/GD/China/2018 shared the highest nucleotide similarity (99.1%) with one wild boar-origin SVV strain (Sichuan HS-01) from the Sichuan province of China. Genetic analysis revealed that SVA/GD/China/2018 clustered with those porcine-origin SVV strains. To the best of our knowledge, this is the first report of SVV infection in buffalo, which might expand the host range of the virus. Surveillance should be expanded, and clinical significance of SVV needs to be further evaluated in cattle.

Use of dual priming oligonucleotide system-based multiplex RT-PCR assay to detect five diarrhea viruses in pig herds in South China.

In this study, a specific and simple method based on the dual priming oligonucleotide (DPO) system was developed to simultaneously detect transmissible gastroenteritis virus (TGEV), porcine epidemic diarrhea virus (PEDV), porcine rotavirus A (PRV-A), porcine delta coronavirus (PDCoV), and swine acute diarrhea syndrome coronavirus (SADS-CoV), associated with the major enteric RNA viruses in pigs. The DPO system-based multiplex RT-PCR method simplified the primer design and did not require optimization of the annealing temperature. Specificity analysis revealed that the method could specifically detect TGEV, PEDV, PRV-A, PDCoV, and SADS-CoV without any cross-amplification of other circulating swine viruses. The limit of detection of the method was as low as 103-104 copies/µL plasmid of each virus. The method also had good repeatability, and obvious results were seen in three repeat experiments with an interval of 45 days. This optimized multiplex RT-PCR method was used to evaluate 181 clinical swine samples that were collected from four provinces of China between September 2016 and August 2018. The results showed that the positive detection rates of PEDV, PDCoV, SADS-CoV, PRV-A, and TGEV were 30.94% (56/181), 17.67% (32/181), 11.6% (21/181), 9.39% (17/181), and 0.55% (1/181), respectively. Mixed infection of two or more viruses was also common. The DPO system-based multiplex RT-PCR could be a useful tool for detecting enteric virus infections. This method has the advantages of easy operation, low cost, high detection efficiency, and short running time for early diagnosis in clinical cases.

The Prevalence, Genetic Characterization, and Evolutionary Analysis of Porcine Pegivirus in Guangdong, China.

Porcine pegivirus (PPgV) is a member of the Pegivirus genus in the Flaviviridae family. PPgV is an emerging virus that has been discovered in swine herds in Germany, the United States, China, Poland, Italy, and the United Kingdom, indicating a wide geographical distribution. In this retrospective study, 339 pig serum samples were collected from 20 different commercial swine farms located in nine cities in Guangdong Province, China, from 2016 to 2018, to investigate the prevalence and genetic diversity of PPgV in this geographical region. PPgV was detected in 55% (11/20) of the farms using nested reverse transcription PCR, with 6.2% (21/339) of pigs testing positive for PPgV. The yearly PPgV-positive rate increased from 2.6% to 7.5% between 2016 and 2018. Sequencing of PPgV-positive samples identified two complete polyprotein genes and seven partial NS5B genes from different farms. Comparative analysis of the polyprotein genes revealed that PPgV sequences obtained in this study showed 87.4%-97.2% similarity at the nucleotide level and 96.5%-99.4% similarity at the amino acid level with the reference sequences. Sequence alignment and phylogenetic analysis of the complete polyprotein gene and partial NS5B and NS3 genes demonstrated a high genetic similarity with the samples from the USA. The finding of the wide distribution of PPgV in swine herds in Guangdong Province will contribute to the understanding of the epidemiological characteristics and genetic evolution of PPgV in China.

Rapid and sensitive detection of Senecavirus A by reverse transcription loop-mediated isothermal amplification combined with a lateral flow dipstick method.

Senecavirus A (SVA) is a critical pathogen causing vesicular lesions in sows and acute death of newborn piglets, resulting in very large economic losses in the pig industry. To restrict the transmission of SVA, an establishment of an effective diagnostic method is crucial for the prevention and control of the disease. However, traditional detection methods often have many drawbacks. In this study, reverse transcription loop-mediated isothermal amplification (RT-LAMP) was combined with a lateral flow dipstick (LFD) to detect SVA. The resulting RT-LAMP-LFD assay was performed at 60°C for 50 min and then directly judged on an LFD visualization strip. This method shows high specificity and sensitivity to SVA. The detection limit of RT-LAMP was 4.56x10-8 ng/µL RNA, approximately 11 copies/µL RNA, and it was 10 times more sensitive than RT-PCR. This detection method's positive rate for clinical samples is comparable to that of RT-PCR. This method is time saving and highly efficient and is thus expected to be used to diagnose SVA infections in this field.