Attenuated SARS-CoV-2 variants with deletions at the S1/S2 junction
EMERGING MICROBES & INFECTIONS
Authors: Lau, Siu-Ying; Wang, Pui; Mok, Bobo Wing-Yee; Zhang, Anna Jinxia; Chu, Hin; Lee, Andrew Chak-Yiu; Deng, Shaofeng; Chen, Pin; Chan, Kwok-Hung; Song, Wenjun; Chen, Zhiwei; To, Kelvin Kai-Wang; Chan, Jasper Fuk-Woo; Yuen, Kwok-Yung; Chen, Honglin
The emergence of SARS-CoV-2 has led to the current global coronavirus pandemic and more than one million infections since December 2019. The exact origin of SARS-CoV-2 remains elusive, but the presence of a distinct motif in the S1/S2 junction region suggests the possible acquisition of cleavage site(s) in the spike protein that promoted cross-species transmission. Through plaque purification of Vero-E6 cultured SARS-CoV-2, we found a series of variants which contain 15-30-bp deletions (Del-mut) or point mutations respectively at the S1/S2 junction. Examination of the original clinical specimen from which the isolate was derived, and 26 additional SARS-CoV-2 positive clinical specimens, failed to detect these variants. Infection of hamsters shows that one of the variants (Del-mut-1) which carries deletion of 10 amino acids (30bp) does not cause the body weight loss or more severe pathological changes in the lungs that is associated with wild type virus infection. We suggest that the unique cleavage motif promoting SARS-CoV-2 infection in humans may be under strong selective pressure, given that replication in permissive Vero-E6 cells leads to the loss of this adaptive function. It would be important to screen the prevalence of these variants in asymptomatic infected cases. The potential of the Del-mut variants as an attenuated vaccine or laboratory tool should be evaluated.
Efficient production of recombinant SARS-CoV-2 spike protein using the baculovirus-silkworm system
BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS
Authors: Fujita, Ryosuke; Hino, Masato; Ebihara, Takeru; Nagasato, Takumi; Masuda, Akitsu; Lee, Jae Man; Fujii, Tsuguru; Mon, Hiroaki; Kakino, Kohei; Nagai, Ryo; Tanaka, Miyu; Tonooka, Yoshino; Moriyama, Takato; Kusakabe, Takahiro
In the case of a new viral disease outbreak, an immediate development of virus detection kits and vaccines is required. For COVID-19, we established a rapid production procedure for SARS-CoV-2 spike protein (S protein) by using the baculovirus-silkworm expression system. The baculovirus vector-derived S proteins were successfully secreted to silkworm serum, whereas those formed insoluble structure in the larval fat body and the pupal cells. The ectodomain of S protein with the native sequence was cleaved by the host furin-protease, resulting in less recombinant protein production. The S protein modified in furin protease-target site was efficiently secreted to silkworm serum and was purified as oligomers, which showed immunoreactivity for anti-SARS-CoV-2 S2 antibody. By using the direct transfection of recombinant bacmid to silkworms, we achieved the efficient production of SARS-CoV-2 S protein as fetal bovine serum (FBS)-free system. The resultant purified S protein would be useful tools for the development of immunodetection kits, antigen for immunization for immunoglobulin production, and vaccines. (C) 2020 Published by Elsevier Inc.