tetano
Editor, Senior Moderator
J Clin Virol Plus
. 2021 Jun;1(1):100012.
doi: 10.1016/j.jcvp.2021.100012. Epub 2021 Apr 5.
Comparative performances of seven quantitative Reverse-Transcription Polymerase Chain Reaction assays (RT-qPCR) for detecting SARS-CoV-2 infection in samples from individuals suspected of COVID-19 in São Paulo, Brazil
Lucila Okuyama Fukasawa[SUP] 1 [/SUP], Cláudio Tavares Sacchi[SUP] 2 [/SUP], Maria Gisele Gonçalves[SUP] 1 [/SUP], Ana Paula Silva Lemos[SUP] 3 [/SUP], Samanta Cristine Grassi Almeida[SUP] 3 [/SUP], Adele Caterino-de-Araujo[SUP] 1 [/SUP]
Affiliations
Abstract
Introduction: Brazil is the second largest country with COVID-19 positive cases worldwide. Due to the potent spread of the virus and the scarcity of kits and supplies, the Brazilian Ministry of Health has granted authorization for the use of kits available during this emergency, without an accurate evaluation of their performance. This study compared the performance and cost-effectiveness of seven molecular assays/kits available in São Paulo, Brazil, for SARS-CoV-2 diagnosis.
Materials and methods: A total of 205 nasopharyngeal/oropharyngeal samples from suspected cases of COVID-19, were tested using the following assays: (i) GeneFinder COVID-19 plus RealAmp kit; (ii) 2019-nCoV RNA PCR-Fluorescence Probing, Da An Gene Co.; (iii) in-house RT-qPCR SARS-CoV-2 IAL; (iv) 2019-nCoV kit, IDT; (v) molecular SARS-CoV-2 (E) kit, Bio-Manguinhos; (vi) Allplex 2019-nCoV modified Assay, Seegene Inc, and (vii) Biomol one-step COVID-19 kit, IBMP. The criteria for determining a SARS-CoV-2 true positive result included the cycle threshold cut-off values, the characteristics of exponential/linear curves, the gene target diversity, and a positive result in at least two assays.
Results: The overall sensitivity of the assays listed were GeneFinder 83.6%, Da An Gene 100.0%, IAL 90.4%, IDT 94.6%, Bio-Manguinhos 87.7%, Allplex 97.3%, and IBMP 87.7%. The minor sensitive gene target was RdRP. Although all assays had a Cohen's Kappa index ≥0.893, the best tests used multiplex assays identifying N-gene and/or E-gene targets.
Conclusion: All assays tested accurate for diagnosis, but considering cost-effectiveness (cost, time consumption, number of samples tested, and performance), the in-house IAL assay was ideal for COVID-19 diagnosis in São Paulo, Brazil.
Keywords: Assay performances; CDC, center for disease control; CI, confidence interval; COVID-19, Coronavirus disease 2019; CV, coefficient of variation in percentage; Coronavirus disease 2019 (COVID-19); Cost-effectiveness; Ct, cycle threshold; Cut-off, Ct limit of positivity; Cy5, Cyanine 5; Diagnosis; E, envelope; FAM, fluorescein amidite; HE, hemagglutinin esterase; HEX, hexachloro fluorescein; IAL, Instituto Adolfo Lutz; IC, internal control; M, membrane; MERS-CoV, Middle East respiratory syndrome; N, nucleocapsid; ORF1ab, open-reading frame of ORF1ab region; PAHO, Pan American Health Organization; ROX, carboxy-X-rhodamine; RP, human ribonuclease p; RT-qPCR, reverse transcription-quantitative polymerase chain reaction; RdRP, RNA-dependent RNA polymerase; Reverse transcription-quantitative polymerase chain reaction (RT-qPCR); S, spike; SARS, severe acute respiratory syndrome; SARS-CoV-2; SD, standard deviation; Severe acute respiratory syndrome (SARS); VIC, 2′-chloro-7′-phenyl-1,4-dichloro-6-carboxyfluorescein; WHO, World Health Organization.
. 2021 Jun;1(1):100012.
doi: 10.1016/j.jcvp.2021.100012. Epub 2021 Apr 5.
Comparative performances of seven quantitative Reverse-Transcription Polymerase Chain Reaction assays (RT-qPCR) for detecting SARS-CoV-2 infection in samples from individuals suspected of COVID-19 in São Paulo, Brazil
Lucila Okuyama Fukasawa[SUP] 1 [/SUP], Cláudio Tavares Sacchi[SUP] 2 [/SUP], Maria Gisele Gonçalves[SUP] 1 [/SUP], Ana Paula Silva Lemos[SUP] 3 [/SUP], Samanta Cristine Grassi Almeida[SUP] 3 [/SUP], Adele Caterino-de-Araujo[SUP] 1 [/SUP]
Affiliations
- PMID: 35262000
- PMCID: PMC8019594
- DOI: 10.1016/j.jcvp.2021.100012
Abstract
Introduction: Brazil is the second largest country with COVID-19 positive cases worldwide. Due to the potent spread of the virus and the scarcity of kits and supplies, the Brazilian Ministry of Health has granted authorization for the use of kits available during this emergency, without an accurate evaluation of their performance. This study compared the performance and cost-effectiveness of seven molecular assays/kits available in São Paulo, Brazil, for SARS-CoV-2 diagnosis.
Materials and methods: A total of 205 nasopharyngeal/oropharyngeal samples from suspected cases of COVID-19, were tested using the following assays: (i) GeneFinder COVID-19 plus RealAmp kit; (ii) 2019-nCoV RNA PCR-Fluorescence Probing, Da An Gene Co.; (iii) in-house RT-qPCR SARS-CoV-2 IAL; (iv) 2019-nCoV kit, IDT; (v) molecular SARS-CoV-2 (E) kit, Bio-Manguinhos; (vi) Allplex 2019-nCoV modified Assay, Seegene Inc, and (vii) Biomol one-step COVID-19 kit, IBMP. The criteria for determining a SARS-CoV-2 true positive result included the cycle threshold cut-off values, the characteristics of exponential/linear curves, the gene target diversity, and a positive result in at least two assays.
Results: The overall sensitivity of the assays listed were GeneFinder 83.6%, Da An Gene 100.0%, IAL 90.4%, IDT 94.6%, Bio-Manguinhos 87.7%, Allplex 97.3%, and IBMP 87.7%. The minor sensitive gene target was RdRP. Although all assays had a Cohen's Kappa index ≥0.893, the best tests used multiplex assays identifying N-gene and/or E-gene targets.
Conclusion: All assays tested accurate for diagnosis, but considering cost-effectiveness (cost, time consumption, number of samples tested, and performance), the in-house IAL assay was ideal for COVID-19 diagnosis in São Paulo, Brazil.
Keywords: Assay performances; CDC, center for disease control; CI, confidence interval; COVID-19, Coronavirus disease 2019; CV, coefficient of variation in percentage; Coronavirus disease 2019 (COVID-19); Cost-effectiveness; Ct, cycle threshold; Cut-off, Ct limit of positivity; Cy5, Cyanine 5; Diagnosis; E, envelope; FAM, fluorescein amidite; HE, hemagglutinin esterase; HEX, hexachloro fluorescein; IAL, Instituto Adolfo Lutz; IC, internal control; M, membrane; MERS-CoV, Middle East respiratory syndrome; N, nucleocapsid; ORF1ab, open-reading frame of ORF1ab region; PAHO, Pan American Health Organization; ROX, carboxy-X-rhodamine; RP, human ribonuclease p; RT-qPCR, reverse transcription-quantitative polymerase chain reaction; RdRP, RNA-dependent RNA polymerase; Reverse transcription-quantitative polymerase chain reaction (RT-qPCR); S, spike; SARS, severe acute respiratory syndrome; SARS-CoV-2; SD, standard deviation; Severe acute respiratory syndrome (SARS); VIC, 2′-chloro-7′-phenyl-1,4-dichloro-6-carboxyfluorescein; WHO, World Health Organization.