Clinical evaluation of a fully automated and high-throughput molecular testing system for detection of influenza virus

Kosai, Kosuke; Kaku, Norihito; Horie, Michiko; Kodama, Hina; Akamatsu, Norihiko; Narita, Yusuke; Matsumoto, Yasushi; Matsushita, Takayuki; Mizuta, Yohei; Izumikawa, Koichi; Mukae, Hiroshi; Yanagihara, Katsunori

doi:10.1186/s12985-022-01916-w

Cited by 1 publication

(1 citation statement)

References 8 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Reverse transcription-polymerase chain reaction (RT-PCR) is a widely used method that provides high sensitivity, specificity, and good repeatability ( Jokela et al, 2010 ; Tang et al, 2020 ; Zhang et al, 2020 ). To date, several commercial detection kits based on RT-PCR have been developed and used, such as the CDC’s Influenza SARS-CoV-2 (Flu SC2) Multiplex Assay ( Shu et al, 2021 ), the Roche Cobas ® SARS-CoV-2 and Influenza A/B Assay ( Kosai et al, 2022 ), and Cepheid’s Xpert Xpress SARS-CoV-2/Flu/RSV ( Leung et al, 2021 ). The use of multiple detection methods based on RT-PCR mainly depends on two detection modes.…”

Section: Introductionmentioning

confidence: 99%

A novel assay based on DNA melting temperature for multiplexed identification of SARS-CoV-2 and influenza A/B viruses

Gao,

Fan,

Kong

et al. 2023

Front. Microbiol.

View full text Add to dashboard Cite

IntroductionThe severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) and influenza viruses can cause respiratory illnesses with similar clinical symptoms, making their differential diagnoses challenging. Additionally, in critically ill SARS-CoV-2–infected patients, co-infections with other respiratory pathogens can lead to severe cytokine storm and serious complications. Therefore, a method for simultaneous detection of SARS-CoV-2 and influenza A and B viruses will be clinically beneficial.MethodsWe designed an assay to detect five gene targets simultaneously via asymmetric PCR-mediated melting curve analysis in a single tube. We used specific probes that hybridize to corresponding single-stranded amplicons at low temperature and dissociate at high temperature, creating different detection peaks representing the targets. The entire reaction was conducted in a closed tube, which minimizes the risk of contamination. The limit of detection, specificity, precision, and accuracy were determined.ResultsThe assay exhibited a limit of detection of <20 copies/μL for SARS-CoV-2 and influenza A and <30 copies/μL for influenza B, with high reliability as demonstrated by a coefficient of variation for melting temperature of <1.16% across three virus concentrations. The performance of our developed assay and the pre-determined assay showed excellent agreement for clinical samples, with kappa coefficients ranging from 0.98 (for influenza A) to 1.00 (for SARS-CoV-2 and influenza B). No false-positive, and no cross-reactivity was observed with six common non-influenza respiratory viruses.ConclusionThe newly developed assay offers a straightforward, cost-effective and nucleic acid contamination-free approach for simultaneous detection of the SARS-CoV-2, influenza A, and influenza B viruses. The method offers high analytical sensitivity, reliability, specificity, and accuracy. Its use will streamline testing for co-infections, increase testing throughput, and improve laboratory efficacy.

show abstract