Back to Science in Searching for SARS-CoV-2 Origins

**Background:** The origin of SARS-CoV-2 remains unknown after more than three years of pandemic. The authors, from the Chinese Center for Disease Control and Prevention, argue that origin tracing is systematic, time-consuming work requiring rigorous logical relationships, sufficient causal relationships, and a solid chain of evidence. They distinguish between the global origin of COVID-19 (likely zoonotic) and the narrower origin or introduction of the index case in a particular geographic area. The paper reviews evidence from environmental and animal sampling at the Huanan Seafood Market (HSM) in Wuhan, China, which was linked to early cases in late December 2019.

**Methods:** The HSM was closed on January 1, 2020. From January to March 2020, Chinese CDC and local CDCs collected environmental samples from locations within and around the market, as well as animal samples including animal bodies, stray animals, and their feces. A total of 1,380 samples were collected. Samples were tested by SARS-CoV-2-specific RT-qPCR. Positive samples underwent virus isolation and RNA-seq analysis. DNA barcoding analysis was performed to identify vertebrate species present in environmental samples. Raw data were deposited in GISAID, SRA/NCBI, NGDC/CNCB, and NMDC.

**Key Results:** Of the 1,380 samples, 73 environmental samples tested positive for SARS-CoV-2 by RT-qPCR, and three live viruses were successfully isolated. The viruses from the market shared nucleotide identity of 99.980% to 100% with SARS-CoV-2 isolates from early COVID-19 cases. No virus was detected in animal swabs covering 18 species of animals in the market. RNA-seq analysis of positive and negative environmental samples showed the abundance of different vertebrate species. DNA barcoding analysis revealed high abundance of *Homo sapiens* in environmental samples, suggesting that SARS-CoV-2 contamination derived from early human cases in the market. The analysis also detected *Sus*, *Bos*, *Gallus*, *Anas*, and *Nyctereutes* (raccoon dog) DNA in environmental samples, but these findings could not prove infection of animals, and human-to-animal transmission could not be ruled out. The authors conclude that the market acted as an early amplifier during the pandemic due to high consumer traffic, causing many initially identified infection clusters.

**Clinical Implications:** The authors argue that based on current scientific logic and globally available evidence, the hypotheses and conclusions on SARS-CoV-2 origins in the previous WHO-China Joint Report remain scientific and objective. They emphasize that traceability is a scientific issue requiring scientific evidence and logical inference, and that the origin of SARS-CoV-2 may remain unknown for a long time. The paper highlights that even if the origin within a certain area is found, it cannot be equated with the origin of the global pandemic—'the location of discovery' does not equal 'the location of origin.' The authors call for continued scientific investigation and note that experience from COVID-19 studies will inform prevention and control of future emerging infectious diseases.