Total infectome characterization of respiratory infections in pre-COVID-19 Wuhan, China

At the end of 2019 Wuhan witnessed an outbreak of "atypical pneumonia" that later developed into a global pandemic. Metagenomic sequencing rapidly revealed the causative agent of this outbreak to be a novel coronavirus denoted SARS-CoV-2. To provide a snapshot of the pathogens in pneumonia-associated respiratory samples from Wuhan prior to the emergence of SARS-CoV-2, we collected bronchoalveolar lavage fluid samples from 408 patients presenting with pneumonia and acute respiratory infections at the Central Hospital of Wuhan between 2016 and 2017. Unbiased total RNA sequencing was performed to reveal their "total infectome", including viruses, bacteria and fungi. We identified 35 pathogen species, comprising 13 RNA viruses, 3 DNA viruses, 16 bacteria and 3 fungi, often at high abundance and including multiple co-infections (13.5%). SARS-CoV-2 was not present. These data depict a stable core infectome comprising common respiratory pathogens such as rhinoviruses and influenza viruses, an atypical respiratory virus (EV-D68), and a single case of a sporadic zoonotic pathogen-Chlamydia psittaci. Samples from patients experiencing respiratory disease on average had higher pathogen abundance than healthy controls. Phylogenetic analyses of individual pathogens revealed multiple origins and global transmission histories, highlighting the connectedness of the Wuhan population. This study provides a comprehensive overview of the pathogens associated with acute respiratory infections and pneumonia, which were more diverse and complex than obtained using targeted PCR or qPCR approaches. These data also suggest that SARS-CoV-2 or closely related viruses were absent from Wuhan in 2016-2017. Author summaryAcute respiratory infections and pneumonia are a significant public health concern on a global scale. This was brought sharply into focus by the emergence of COVID-19 at the end of 2019 in Wuhan, China. Far less is known, however, about the total "infectomes" associated with respiratory infection and pneumonia: that is, all the viruses, bacteria and fungi that might be associated with these important human diseases. Herein, we describe total infectomes in bronchoalveolar lavage fluid samples taken from 408 patients presenting with pneumonia and acute respiratory infection in a single hospital in Wuhan between 2016 and 2017. From these samples we identified 35 pathogen species, comprising 13 RNA viruses, 3 DNA viruses, 16 bacteria and 3 fungi, often at high abundance and including multiple co-infections. We found no evidence for SARS-CoV-2. These data reveal a stable core infectome comprising common respiratory pathogens as well as their history of global transmission.