Ultrasensitive Hexaplex Droplet Digital Polymerase Chain Reaction Assay for Rapid Screening and Quantification of Genetically Modified Content

2021 ◽  
Author(s):  
Rong Li ◽  
Wenting Xu ◽  
Xiujie Zhang ◽  
Xueqi Li ◽  
Jinjie Cui ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Polymerase Chain Reaction Assay ◽  
Genetically Modified ◽  
Rapid Screening ◽  
Chain Reaction ◽  
Polymerase Chain ◽  
Digital Polymerase Chain Reaction

scholarly journals Natural Transmission of Bat-like Severe Acute Respiratory Syndrome Coronavirus 2 Without Proline-Arginine-Arginine-Alanine Variants in Coronavirus Disease 2019 Patients

2020 ◽  
Author(s):  
Yik Chun Wong ◽  
Siu Ying Lau ◽  
Kelvin Kai Wang To ◽  
Bobo Wing Yee Mok ◽  
Xin Li ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Severe Acute Respiratory Syndrome ◽  
Prevalence Rate ◽  
Polymerase Chain Reaction Assay ◽  
Wild Type ◽  
Natural Evolution ◽  
Chain Reaction ◽  
Furin Cleavage ◽  
Polymerase Chain ◽  
Digital Polymerase Chain Reaction

Abstract Background Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) contains the furin cleavage Proline-Arginine-Arginine-Alanine (PRRA) motif in the S1/S2 region, which enhances viral pathogenicity but is absent in closely related bat and pangolin coronaviruses. Whether bat-like coronaviral variants without PRRA (∆PRRA) can establish natural infections in humans is unknown. Methods Here, we developed a duplex digital polymerase chain reaction assay to examine ∆PRRA variants in Vero-E6-propagated isolates, human organoids, experimentally infected hamsters, and coronavirus disease 2019 (COVID-19) patients. Results We found that SARS-CoV-2, as currently transmitting in humans, contained a quasispecies of wild-type, ∆PRRA variants and variants that have mutations upstream of the PRRA motif. Moreover, the ∆PRRA variants were readily detected despite being at a low intra-host frequency in transmitted founder viruses in hamsters and in COVID-19 patients, including in acute cases and a family cluster, with a prevalence rate of 52.9%. Conclusions Our findings demonstrate that bat-like SARS-CoV-2ΔPRRA not only naturally exists but remains transmissible in COVID-19 patients, which has significant implications regarding the zoonotic origin and natural evolution of SARS-CoV-2.

Sign in / Sign up

Export Citation Format

Share Document