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Mem Inst Oswaldo Cruz, Rio de Janeiro, VOLUME 119 | 2024

Research Articles

Point-of-care testing for COVID-19: a simple two-step molecular diagnostic development and validation during the SARS-CoV-2 pandemic

Andre Akira Gonzaga Yoshikawa¹, Sabrina Fernandes Cardoso^1,2, Lívia Budziarek Eslabão³, Iara Carolini Pinheiro¹, Priscila Valverde⁴, Gisele Caminha⁵, Oscar Bruna Romero³, Leandro Medeiros⁶, Luísa Damazio Pitaluga Rona^1,7,+, André Nóbrega Pitaluga^7,8,+

¹Universidade Federal de Santa Catarina, Departamento de Biologia Celular, Embriologia e Genética, Florianópolis, SC, Brasil
²Secretaria de Saúde do Estado de Santa Catarina, Diretoria de Vigilância Epidemiológica, Florianópolis, SC, Brasil
³Universidade Federal de Santa Catarina, Departamento de Microbiologia, Imunologia e Parasitologia, Florianópolis, SC, Brasil
⁴Secretaria Municipal de Saúde, Florianópolis, SC, Brasil
⁵Laboratório Central de Saúde Pública de Santa Catarina, Florianópolis, SC, Brasil
⁶Instituto Federal de Educação, Ciência e Tecnologia de Santa Catarina, Florianópolis, SC, Brasil
⁷Conselho Nacional de Desenvolvimento Científico e Tecnológico, Instituto Nacional de Ciência e Tecnologia em Entomologia Molecular, Rio de Janeiro, RJ, Brasil
⁸Fundação Oswaldo Cruz-Fiocruz, Instituto Oswaldo Cruz, Rio de Janeiro, RJ, Brasil

DOI: 10.1590/0074-02760230236

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ABSTRACT

BACKGROUND During the coronavirus disease 19 (COVID-19) pandemic, diagnostic testing of the general population proved challenging due to limitations of the gold-standard diagnostic procedure using reverse transcription real-time polymerase chain reaction (RT-qPCR) for large-scale testing on the centralised model, especially in low-resource areas.
OBJECTIVES To address this, a point-of-care (PoC) diagnostic protocol for COVID-19 was developed, providing fast, reliable, and affordable testing, particularly for low-mid develop areas.
METHODS The PoC diagnostic process combines a simple paper-based RNA extraction method housed within a 3D-printed plastic device with a colorimetric reverse transcription loop-mediated isothermal amplification (RT-LAMP) assay. Nasopharyngeal/oropharyngeal swabs (NOS) and saliva samples were tested between 2020 and 2021, with the assistance of Santa Catarina’s State Health Secretary, Brazil.
FINDINGS The developed diagnostic protocol showed a limit of detection of 9,900 copies and an overall diagnostic specificity of 98% for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) from 1,348 clinical analysed samples. The diagnostic sensitivity was 95% for NOS samples, 85% for early morning saliva, and 69% for indiscriminate saliva.
MAIN CONCLUSIONS In conclusion, the developed device successfully extracted SARS-CoV-2 viral RNA from swabs and saliva clinical samples. When combined with colorimetric RT-LAMP, it provides results within 45 min using minimal resources, thus delivering a diagnostic kit protocol that is applicable in large-scale sampling.

key words: COVID-19 SARS-CoV-2 point-of-care molecular diagnostic RT-LAMP

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