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Natural transmission of bat-like SARS-CoV-2 △PRRA variants in COVID-19 patients

Our take —

In a study of 51 COVID-19 patients from Hong Kong, researchers detected SARS-CoV-2 variants with deletions in and around the spike S1/S2 cleavage site, a feature shared with related viruses in bats and pangolins. These results, combined with results showing S1/S2 site insertions in related bat coronaviruses, indicate that the PRRA motif of SARS-CoV-2 is genetically labile but has a selective advantage over variants in experimental models and infected humans. These results suggest that natural selection (occurring in animals prior to spillover, or in humans shortly after spillover) has fixed the PRRA insertion variant in SARS-CoV-2. Nevertheless, the exact origin of the PRRA motif in SARS-CoV-2 from an animal virus has not yet been determined and will require further sampling to clarify the selective processes that have shaped SARS-CoV-2.

Study design

Ecological, Other

Study population and setting

SARS-CoV-2 contains a unique furin cleavage site amino acid insertion (PRRA) in the S1/S2 region of the spike protein that is absent in related coronaviruses in bats and pangolins. The authors developed a PCR-based assay to detect viral variants with deletions of the PRRA site or upstream of the site and distinguish these variants from normal (wildtype) SARS-CoV-2. The infectivity of wildtype and S1/S2 cleavage site variants was tested in human intestinal cell clusters (organoids; n = 3) and hamsters (n = 2). The presence of viral variants in human COVID-19 patients was assessed in clinical specimens (saliva, nasopharyngeal secretions, throat swabs, and endotracheal aspirates) from 51 adult patients from Hong Kong, China.

Summary of Main Findings

The authors determined that their assay was capable of detecting S1/S2 cleavage site variants at low amounts, down to single viral copies. Inoculation of human intestinal organoids and hamsters with a wildtype virus containing a low copy number of viral variants with PRRA deletions and upstream deletions resulted in infections, although the copies of cleavage site variants were suppressed relative to the inoculum. In the samples from 51 COVID-19 patients, wildtype viral sequences were the most abundant (99% of viral molecules on average). A majority of samples (53%) had PRRA deletion variant molecules, but the mutant population was always a minority of molecules (0.3% on average). Upstream variants were observed in 82% of samples and the mutant represented 1% of detected viral copies. Upstream variants were more common than deletion variants in 93% of samples. These results suggest that cleavage site variants are less competitive than the wildtype virus with the PRRA insertion in cell culture and in natural human infections. The presence of variants had no relationship with symptom severity or sample type; upstream variants were always more common than PRRA deletion variants.

Study Strengths

The authors developed a sensitive assay to detect SARS-CoV-2 genetic variants in clinical samples. Cell culture and a hamster animal model were useful in determining the relative replication fitness of variants compared to wildtype SARS-CoV-2.

Limitations

The study size was very limited, including only 51 patients, three intestinal organoids, and two hamsters. Human samples only came from adult patients in Hong Kong, so the presence of S1/S2 cleavage site variants in other geographic areas cannot be determined from this study. The study also cannot determine how SARS-CoV-2 acquired its unique PRRA motif, or whether that event took place in humans or occurred in an animal host prior to spillover into humans.

Value added

The study provided evidence that natural spike S1/S2 cleavage site variants of SARS-CoV-2, similar to related viruses in bats and pangolins, are currently circulating in humans with COVID-19; however, the low frequency of these variants in natural infections indicates that they are less competitive than the wildtype with the PRRA insertion.

This review was posted on: 22 July 2020