SARS-CoV-2 antigens expressed in plants detect antibody responses in COVID-19 patients. Makatsa, M. S, Tincho, M. B, Wendoh, J. M, Ismail, S. D, Nesamari, R., Pera, F., de Beer, S., David, A., Jugwanth, S., Gededzha, M. P, Mampeule, N., Sanne, I., Stevens, W., Scott, L., Blackburn, J., Mayne, E. S, Keeton, R. S, & Burgers, W. A medRxiv, jan, 2020.
SARS-CoV-2 antigens expressed in plants detect antibody responses in COVID-19 patients [link]Paper  doi  abstract   bibtex   
Background: The SARS-CoV-2 pandemic has swept the world and poses a significant global threat to lives and livelihoods, with over 16 million confirmed cases and at least 650 000 deaths from COVID-19 in the first 7 months of the pandemic. Developing tools to measure seroprevalence and understand protective immunity to SARS-CoV-2 is a priority. We aimed to develop a serological assay using plant-derived recombinant viral proteins, which represent important tools in less-resourced settings. Methods: We established an indirect enzyme-linked immunosorbent assay (ELISA) using the S1 and receptor-binding domain (RBD) portions of the spike protein from SARS-CoV-2, expressed in Nicotiana benthamiana. We measured antibody responses in sera from South African patients (n=77) who had tested positive by PCR for SARS-CoV-2. Samples were taken a median of six weeks after the diagnosis, and the majority of participants had mild and moderate COVID-19 disease. In addition, we tested the reactivity of pre-pandemic plasma (n=58) and compared the performance of our in-house ELISA with a commercial assay. We also determined whether our assay could detect SARS-CoV-2-specific IgG and IgA in saliva. Results: We demonstrate that SARS-CoV-2-specific immunoglobulins are readily detectable using recombinant plant-derived viral proteins, in patients who tested positive for SARS-CoV-2 by PCR. Reactivity to S1 and RBD was detected in 51 (66%) and 48 (62%) of participants, respectively. Notably, we detected 100% of samples identified as having S1-specific antibodies by a validated, high sensitivity commercial ELISA, and OD values were strongly and significantly correlated between the two assays. For the pre-pandemic plasma, 1/58 (1.7%) of samples were positive, indicating a high specificity for SARS-CoV-2 in our ELISA. SARS-CoV-2-specific IgG correlated significantly with IgA and IgM responses. Endpoint titers of S1- and RBD-specific immunoglobulins ranged from 1:50 to 1:3200. S1-specific IgG and IgA were found in saliva samples from convalescent volunteers. Conclusions: We demonstrate that recombinant SARS-CoV-2 proteins produced in plants enable robust detection of SARS-CoV-2 humoral responses. This assay can be used for seroepidemiological studies and to measure the strength and durability of antibody responses to SARS-CoV-2 in infected patients in our setting.Competing Interest StatementFrancisco Pera and Scott de Beer are employed by Cape Bio Pharms. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.Funding StatementThis work was supported by the Wellcome Centre for Infectious Diseases Research in Africa (CIDRI-Africa), which is supported by core funding from the Wellcome Trust [203135/Z/16/Z]. Sample collection was funded through the EQUIP grant AID-OAA-A-15-00070- Antiretroviral Therapy Simplification-Optimization of Programs and Services (ART-OPS) COVID supplement and through iLEAD BMGF (i-LEAD) grant ID OPP1171455. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. The views expressed are those of the authors, and the funders are not responsible for any use that may be made of the information contained herein. Author DeclarationsI confirm all relevant ethical guidelines have been followed, and any necessary IRB and/or ethics committee approvals have been obtained.YesThe details of the IRB/oversight body that provided approval or exemption for the research described are given below:Human Research Ethics Committee (HREC) of the University of Witwatersrand (M200468) and the University of Cape Town (UCT; 210/2020).All necessary patient/participant consent has been obtained and the appropriate institutional forms have been archived.YesI understand that all clinical trials and any other prospective interventional studies must be registered with an ICMJE-approved registry, such as ClinicalTrials.gov. I confirm that any such study reported in the manuscript has been registered and the trial registration ID is provided (note: if posting a prospective study registered retrospectively, please provide a statement in the trial ID field explaining why the study was not registered in advance).Yes I have followed all appropriate research reporting guidelines and uploaded the relevant EQUATOR Network research reporting checklist(s) and other pertinent material as supplementary files, if applicable.YesAll data is available and can be obtained by contacting the corresponding author.
@article{Makatsa2020,
abstract = {Background: The SARS-CoV-2 pandemic has swept the world and poses a significant global threat to lives and livelihoods, with over 16 million confirmed cases and at least 650 000 deaths from COVID-19 in the first 7 months of the pandemic. Developing tools to measure seroprevalence and understand protective immunity to SARS-CoV-2 is a priority. We aimed to develop a serological assay using plant-derived recombinant viral proteins, which represent important tools in less-resourced settings. Methods: We established an indirect enzyme-linked immunosorbent assay (ELISA) using the S1 and receptor-binding domain (RBD) portions of the spike protein from SARS-CoV-2, expressed in Nicotiana benthamiana. We measured antibody responses in sera from South African patients (n=77) who had tested positive by PCR for SARS-CoV-2. Samples were taken a median of six weeks after the diagnosis, and the majority of participants had mild and moderate COVID-19 disease. In addition, we tested the reactivity of pre-pandemic plasma (n=58) and compared the performance of our in-house ELISA with a commercial assay. We also determined whether our assay could detect SARS-CoV-2-specific IgG and IgA in saliva. Results: We demonstrate that SARS-CoV-2-specific immunoglobulins are readily detectable using recombinant plant-derived viral proteins, in patients who tested positive for SARS-CoV-2 by PCR. Reactivity to S1 and RBD was detected in 51 (66{\%}) and 48 (62{\%}) of participants, respectively. Notably, we detected 100{\%} of samples identified as having S1-specific antibodies by a validated, high sensitivity commercial ELISA, and OD values were strongly and significantly correlated between the two assays. For the pre-pandemic plasma, 1/58 (1.7{\%}) of samples were positive, indicating a high specificity for SARS-CoV-2 in our ELISA. SARS-CoV-2-specific IgG correlated significantly with IgA and IgM responses. Endpoint titers of S1- and RBD-specific immunoglobulins ranged from 1:50 to 1:3200. S1-specific IgG and IgA were found in saliva samples from convalescent volunteers. Conclusions: We demonstrate that recombinant SARS-CoV-2 proteins produced in plants enable robust detection of SARS-CoV-2 humoral responses. This assay can be used for seroepidemiological studies and to measure the strength and durability of antibody responses to SARS-CoV-2 in infected patients in our setting.Competing Interest StatementFrancisco Pera and Scott de Beer are employed by Cape Bio Pharms. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.Funding StatementThis work was supported by the Wellcome Centre for Infectious Diseases Research in Africa (CIDRI-Africa), which is supported by core funding from the Wellcome Trust [203135/Z/16/Z]. Sample collection was funded through the EQUIP grant AID-OAA-A-15-00070- Antiretroviral Therapy Simplification-Optimization of Programs and Services (ART-OPS) COVID supplement and through iLEAD BMGF (i-LEAD) grant ID OPP1171455. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. The views expressed are those of the authors, and the funders are not responsible for any use that may be made of the information contained herein. Author DeclarationsI confirm all relevant ethical guidelines have been followed, and any necessary IRB and/or ethics committee approvals have been obtained.YesThe details of the IRB/oversight body that provided approval or exemption for the research described are given below:Human Research Ethics Committee (HREC) of the University of Witwatersrand (M200468) and the University of Cape Town (UCT; 210/2020).All necessary patient/participant consent has been obtained and the appropriate institutional forms have been archived.YesI understand that all clinical trials and any other prospective interventional studies must be registered with an ICMJE-approved registry, such as ClinicalTrials.gov. I confirm that any such study reported in the manuscript has been registered and the trial registration ID is provided (note: if posting a prospective study registered retrospectively, please provide a statement in the trial ID field explaining why the study was not registered in advance).Yes I have followed all appropriate research reporting guidelines and uploaded the relevant EQUATOR Network research reporting checklist(s) and other pertinent material as supplementary files, if applicable.YesAll data is available and can be obtained by contacting the corresponding author.},
author = {Makatsa, Mohau S and Tincho, Marius B and Wendoh, Jerome M and Ismail, Sherazaan D and Nesamari, Rofhiwa and Pera, Francisco and de Beer, Scott and David, Anura and Jugwanth, Sarika and Gededzha, Maemu P and Mampeule, Nakampe and Sanne, Ian and Stevens, Wendy and Scott, Lesley and Blackburn, Jonathan and Mayne, Elizabeth S and Keeton, Roanne S and Burgers, Wendy A},
doi = {10.1101/2020.08.04.20167940},
journal = {medRxiv},
keywords = {OA,fund{\_}ack,original},
mendeley-tags = {OA,fund{\_}ack,original},
month = {jan},
pages = {2020.08.04.20167940},
pmid = {33868324},
title = {{SARS-CoV-2 antigens expressed in plants detect antibody responses in COVID-19 patients}},
url = {http://medrxiv.org/content/early/2020/08/04/2020.08.04.20167940.abstract},
year = {2020}
}

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