Highlights d 1.6 million tests identified 1,388 SARS-CoV-2 infections in Guangdong by 19 March d Virus genomes can be recovered using a variety of sequencing approaches d Analyses reveal multiple viral importations with limited local transmission d Effective control measures helped reduce and eliminate chains of viral transmission

No abstract

SARS-CoV-2 is a novel coronavirus first identified in December 2019. Notable features make SARS-CoV-2 distinct from most other previously-identified Betacoronaviruses, including the receptor binding domain of SARS-CoV-2 and a unique insertion of twelve nucleotide or four amino acids (PRRA) at the S1/S2 boundary. In this study, we identified two deletion variants of SARS-CoV-2 that either directly affect the polybasic cleavage site itself (NSPRRAR) or a flanking sequence (QTQTN). These deletions were verified by multiple sequencing methods. In vitro results showed that the deletion of NSPRRAR likely does not affect virus replication in Vero and Vero-E6 cells, however the deletion of QTQTN may restrict late phase viral replication. The deletion of QTQTN was detected in 3 of 68 clinical samples and half of 24 in vitro isolated viruses, whilst the deletion of NSPRRAR was identified in 3 in vitro isolated viruses. Our data indicate that (i) there may be distinct selection pressures on SARS-CoV-2 replication or infection in vitro and in vivo, (ii) an efficient mechanism for deleting this region from the viral genome may exist, given that the deletion variant is commonly detected after two rounds of cell passage, and (iii) the PRRA insertion, which is unique to SARS-CoV-2, is not fixed during virus replication in vitro. These findings provide information to aid further investigation of SARS-CoV-2 infection mechanisms and a better understanding of the NSPRRAR deletion variant observed here. Important notes The spike protein determines the infectivity and host range of coronaviruses. SARS-CoV-2 has two unique features in its spike protein, the receptor binding domain and an insertion of twelve nucleotides at the S1/S2 boundary resulting a furin-like cleavage site. Here, we identified two deletion variants of SARS-CoV-2 that either directly affect the furin-like cleavage site itself (NSPRRAR) or a flanking sequence (QTQTN) and investigated these deletions in cell isolates and clinical samples. The absence of the polybasic cleavage site in SARS-CoV-2 did not affect virus replication in Vero or Vero-E6 cells. Our data indicate the PRRAR and its flanking sites are not fixed in vitro, thus there appears to be distinct selection pressures on SARS-CoV-2 sequences in vitro and in vivo. Further investigation of the mechanism of generating these deletion variants and their infectivity in different animal models would improve our understanding of the origin and evolution of this virus.

BackgroundClonorchis sinensis is a carcinogenic human liver fluke that is widespread in Asian countries. Increasing infection rates of this neglected tropical disease are leading to negative economic and public health consequences in affected regions. Experimental and epidemiological studies have shown a strong association between the incidence of cholangiocarcinoma and the infection rate of C. sinensis. To aid research into this organism, we have sequenced its genome.ResultsWe combined de novo sequencing with computational techniques to provide new information about the biology of this liver fluke. The assembled genome has a total size of 516 Mb with a scaffold N50 length of 42 kb. Approximately 16,000 reliable protein-coding gene models were predicted. Genes for the complete pathways for glycolysis, the Krebs cycle and fatty acid metabolism were found, but key genes involved in fatty acid biosynthesis are missing from the genome, reflecting the parasitic lifestyle of a liver fluke that receives lipids from the bile of its host. We also identified pathogenic molecules that may contribute to liver fluke-induced hepatobiliary diseases. Large proteins such as multifunctional secreted proteases and tegumental proteins were identified as potential targets for the development of drugs and vaccines.ConclusionsThis study provides valuable genomic information about the human liver fluke C. sinensis and adds to our knowledge on the biology of the parasite. The draft genome will serve as a platform to develop new strategies for parasite control.

Highlights d SARS-CoV-2 genome sequencing and phylogenetic analyses identify 35 recurrent mutations d Association with 117 clinical phenotypes reveals potentially important mutations d D500-532 in Nsp1 coding region correlates with lower viral load and serum IFN-b d Viral isolates with D500-532 mutation induce lower IFN-I response in the infected cells