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Volume 17
Issue 12
10.3390/v17121559
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Article

Development and Validation of a Commercial TaqMan-Based RT-qPCR Kit for Rotavirus and Norovirus Detection in the Brazilian Acute Diarrhea Surveillance Network

by
Geison Cambri
1,
Thiago Jacomasso
1,
Fernanda Marcicano Burlandy
2,
Fábio Correia Malta
2,
Alexandre Madi Fialho
2,
Audrey Cilli
3,
Simone Guadagnucci
3,
Dielle Monteiro Teixeira
4,
Patrícia Santos Lobo
4,
Hugo Reis Resque
4,
Lucia Helena Berto
5,
Alessandro Afornali
1,
Irina Nastassja Riediger
1,*,
Luana Silva Soares
4,*,
Rita de Cássia Compagnoli Carmona
3,* and
Tulio Machado Fumian
2,*
1
Instituto de Biologia Molecular do Paraná, Curitiba, PR 81350-010, Brazil
2
Laboratory of Comparative and Environmental Virology (LVCA), Oswaldo Cruz Institute, Oswaldo Cruz Foundation (Fiocruz), Av. Brasil 4365, Rio de Janeiro, RJ 21040-360, Brazil
3
Enteric Diseases Laboratory (LDE), Virology Center (CV), Adolfo Lutz Institute (IAL), Av. Dr Arnaldo, 355, São Paulo, SP 01246-900, Brazil
4
Laboratory of Gastroenteric Virus (LVG), Virology Section, Evandro Chagas Institute, Secretariat of Health Surveillance and Environment, Brazilian Ministry of Health, Ananindeua, PA 67030-000, Brazil
5
General Coordination of Public Health Laboratories (CGLab), Ministry of Health, Brasília, DF 70058-900, Federal District, Brazil
*
Authors to whom correspondence should be addressed.
Viruses 2025, 17(12), 1559; https://doi.org/10.3390/v17121559 (registering DOI)
Submission received: 21 October 2025 / Revised: 24 November 2025 / Accepted: 27 November 2025 / Published: 28 November 2025
(This article belongs to the Special Issue Enteric Viruses in Environment and Humans: Identification, Surveillance and Control)
Versions Notes

Abstract

Acute gastroenteritis (AGE) is a major cause of illness and death in children under five, especially in low- and middle-income countries, and rotavirus A (RVA) and norovirus are the leading viral agents. The present study aimed to describe the development of a commercial multiplex TaqMan-based RT-qPCR assay to detect those viruses to enhance surveillance and public health responses in Brazil. The assay validation involved optimizing primers and probes for multiplex RT-qPCR, assessing analytical sensitivity, and confirming specificity. A multicenter pilot study across Brazil’s AGE surveillance network assessed the assay’s performance. The IBMP NAT assay demonstrated high specificity and sensitivity for detecting RVA and norovirus GI and GII. No cross-reactivity was observed. LoD95 values were low: 18.6 (GI), 71.2 (GII), and 12.3 (RVA) copies/reaction. In 379 clinical samples, diagnostic sensitivity and specificity exceeded 96% for all targets. The assay showed strong reproducibility across operators and instruments. Stability tests confirmed consistent performance under freeze–thaw, transport, and storage conditions. Compared to in-house RT-qPCR, the IBMP NAT test yielded lower Ct values, indicating improved detection of low viral loads. The IBMP NAT Kit significantly advances molecular diagnostics, enabling rapid, sensitive, and reliable detection of RVA and norovirus in fecal specimens. It strengthens public health surveillance and supports timely responses to AGE outbreaks, helping reduce disease burden in vulnerable populations.
Keywords: Acute gastroenteritis; rotavirus; norovirus; diagnostic; quantitative PCR Acute gastroenteritis; rotavirus; norovirus; diagnostic; quantitative PCR

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MDPI and ACS Style

Cambri, G.; Jacomasso, T.; Burlandy, F.M.; Malta, F.C.; Fialho, A.M.; Cilli, A.; Guadagnucci, S.; Teixeira, D.M.; Lobo, P.S.; Resque, H.R.; et al. Development and Validation of a Commercial TaqMan-Based RT-qPCR Kit for Rotavirus and Norovirus Detection in the Brazilian Acute Diarrhea Surveillance Network. Viruses 2025, 17, 1559. https://doi.org/10.3390/v17121559

AMA Style

Cambri G, Jacomasso T, Burlandy FM, Malta FC, Fialho AM, Cilli A, Guadagnucci S, Teixeira DM, Lobo PS, Resque HR, et al. Development and Validation of a Commercial TaqMan-Based RT-qPCR Kit for Rotavirus and Norovirus Detection in the Brazilian Acute Diarrhea Surveillance Network. Viruses. 2025; 17(12):1559. https://doi.org/10.3390/v17121559

Chicago/Turabian Style

Cambri, Geison, Thiago Jacomasso, Fernanda Marcicano Burlandy, Fábio Correia Malta, Alexandre Madi Fialho, Audrey Cilli, Simone Guadagnucci, Dielle Monteiro Teixeira, Patrícia Santos Lobo, Hugo Reis Resque, and et al. 2025. "Development and Validation of a Commercial TaqMan-Based RT-qPCR Kit for Rotavirus and Norovirus Detection in the Brazilian Acute Diarrhea Surveillance Network" Viruses 17, no. 12: 1559. https://doi.org/10.3390/v17121559

APA Style

Cambri, G., Jacomasso, T., Burlandy, F. M., Malta, F. C., Fialho, A. M., Cilli, A., Guadagnucci, S., Teixeira, D. M., Lobo, P. S., Resque, H. R., Berto, L. H., Afornali, A., Riediger, I. N., Soares, L. S., Carmona, R. d. C. C., & Fumian, T. M. (2025). Development and Validation of a Commercial TaqMan-Based RT-qPCR Kit for Rotavirus and Norovirus Detection in the Brazilian Acute Diarrhea Surveillance Network. Viruses, 17(12), 1559. https://doi.org/10.3390/v17121559

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