In 2018, there was an outbreak of African swine fever (ASF) in China, which spread to other provinces in the following three years, severely damaging China’s pig industry. ASF is caused by the African swine fever virus (ASFV). Given that the genome of the African swine fever virus is very complex and whole genome information is currently inadequate, it is important to efficiently obtain virus genome sequences for genomic and epidemiological studies. The prevalent ASFV strains have low genetic variability; therefore, whole genome sequencing analysis provides a basis for the study of ASFV. We provide a method for the efficient sequencing of whole genomes, requiring only a small number of tissues. The pig spleen were crushed and homogenized, and the tissue and cell fragments were removed by low-speed centrifugation. The virus-like particles (VLPs) in the solution were purified and concentrated by filtration and over-speed centrifugation. After obtaining the relatively pure VLP sample, nucleic acids were extracted simultaneously by co-extraction of viral DNA (dsDNA, ssDNA) and RNA (ssRNA, dsRNA). Then, the database construction method was selected according to the genomic types of ASFV, and the whole ASFV genome was obtained through data filtering, host sequence removal, virus classification, data assembly, virus sequence identification, statistical analysis, gene prediction, and functional analysis. Our proposed method will facilitate ASFV genome sequencing and novel virus discovery.

In this study, we detected a circular replication-associated protein (Rep)-encoding single-stranded (CRESS) DNA virus in intestinal tissue samples and faecal samples of pigs. Some researchers named po-circo-like (PCL) virus. PCL virus contains a single-stranded DNA genome, and ORF1 encodes the Rep and not the typical capsid protein encoded in PCV. The Rep protein may be responsible for viral genome replication. In addition, PCL virus may be one of the pathogens causing diarrhea symptoms in pigs. We identified four strains of PCL virus in two different pig farms with severe diarrhea outbreaks in Hunan Province, China. The strains in this study share 39.4%–94.9% nucleic acid identity and 85.3%–98.4% amino acid identity with Rep of the reference strains. A multiple sequence alignment of these PCL viruses and Bo-Circo-like CH showed a identity of 93.2% for the Rep protein, and the nucleotide identity was 86.7-89.3%. Moreover, Bo-Circo-like CH and HN75, HN39-01, HN39-02 had similar stem-loop sequences. The PCL virus might therefore be transmitted to non-porcine hosts by cross-species transmission routes. Through Recombination Detection Program (RDP), Simplot and phylogenetic analyses, strong evidence for recombination events was found in China field PCL virus strains. In conclusion, the present study is the first detailed report of the PCL virus in HuNan provinces, which is a potential new virus in pigs that might be involved in cross-species transmission. Further investigation is needed to determine the pathogenesis of this virus and its epidemiologic impact.