4 Discussion
The outbreak of COVID-19 has raised great concern all over the world and poses serious threat to global public health. Therefore, Covs gained increasing attention. When researchers examined blood samples from Ayiti children in early 2021, three samples were found to be infected with PDCoV (Lednicky et al., 2021). Of note, PDCoV has repeatedly crossed the host barrier to chicken (Boley et al., 2020; Liang et al., 2019). This phenomenon exhibits the same cross-species transmission as SARS-COV-2, which suggested that the emerging porcine delta-coronavirus may be a potential threat to human health. Here, we adopted a phylogeographic approach to examine the time of origin, evolutionary rate, diversification patterns and potential transmission routes of PDCoV and selected six special spike amino acid sequences to align and analyze.
PDCoV is an emerging coronavirus causing severe enteritis, diarrhea, and vomiting in piglets. PDCoV was first discovered in Hong Kong, China in 2012, but began to spread in the United States in 2014. Now many countries in the world have reported that pigs were infected with PDCoV. We attained 42 positive samples for PDCoV from eight intensive pig farms, located in Guangdong Province, China and sequenced two PDCoV completed genomes. Moreover, a strain of PDCoV, which could be stably passaged on the LLC-PK cell line was successfully isolated. According to phylogenetic analysis of PDCoV, our findings indicated that PDCoV would be classified into three major lineages: China lineage; USA lineage; and Southeast Asia lineage. There were three PDCoV in Ayiti that can infect children, two of which were highly similar to KY065120 in Tianjin, China (99.8%), and the rest were highly similar to KR150443 in Arkansas, USA (98.9%). However, after suffering from African swine fever in Ayiti, pigs were reintroduced mainly from American populations. The worldwide spatial dispersal networks of PDCoV also indicated the US lineage was one of the origins to Ayiti. Therefore, PDCoV infected in Ayiti was most likely from America (Alexander, 1992; Lednicky et al., 2021).
The MCC tree showed that PDCoV originated in Asia, between China and southeast Asia, and more likely in Southeast Asia in 1989. In addition, according to Bayesian Skyline Plot analysis, effective population size of PDCoV tended to level off and stabilize at around 70. We observed the association between geographic distance and migration rate to clarify space dynamics. However, we observed a strong signal of viral dissemination (migration rate = 3.55) from US to Japan and South Korea, with a long distance (about 11200 km). This showed there was no certain connection between geographic distance and migration rate. Moreover, the scatter plot of distance and migration rate also did not display a certain trend (Supplementary Tables S6, Fig S2). It may be due to the swine trade between countries and artificial or feed contamination that affects the rate of natural transmission.
To analyse the effect of amino acid mutation on cross-species transmission, we found six PDCoV sequences that can spread across species, and align the amino acids of their spike protein. By analyzing the amino acid sequence of spike protein, seven significant mutation sites were defined. The accumulation of these mutations may enhance dynamic movements, accelerating virus membrane fusion events and transmission.
Frankly, this study has several limitations. First, sampling bias, no same number of samples in different regions, may have affected the Bayesian phylogenic reconstruction and inference of the transmission networks (Alexander, 1992; Lednicky et al., 2021). Second, the information from sequences contains the most regions infected with PDCoV but does not cover all regions, because there are some places infected with PDCoV, but no sequence uploaded in GenBank.
In summary, our findings offer novel insights into the diversification, evolution, and interspecies transmission and origin of PDCoV. We also briefly analyzed why humans are infected with PDCoV. Increasing evidence strongly implicates that CoVs have a high capacity for cross-species transmission. Given that pigs are in frequent contact with humans and wild animals, the lower interspecies hurdles in pigs would make them a potential mixing vessel for CoVs (Ye et al., 2020). Therefore, comprehensive surveillance and enhanced biosecurity precautions for PDCoV should be taken to prevent further pandemic such as COVID-19.