Optimization of a 6-plex Crystal Digital PCR® assay and its application to simultaneous surveillance of enteric and respiratory viruses in wastewater DOI Creative Commons
Bikash Malla, Sadhana Shrestha, Eiji Haramoto

и другие.

The Science of The Total Environment, Год журнала: 2025, Номер 970, С. 178939 - 178939

Опубликована: Март 1, 2025

Multiplex digital PCR (dPCR) approaches are commonly employed in wastewater-based epidemiology (WBE) studies. However, optimizing the dPCR workflow is a critical step to ensure its reliability and accuracy before application. In this study, 6-plex Crystal Digital PCR® (cdPCR) was optimized for simultaneous detection of six epidemiologically important pathogens, including three enteric viruses, noroviruses genogroups I II (NoV-GI GII) enteroviruses (EnV), respiratory severe acute syndrome coronavirus 2 (SARS-CoV-2), which causative agent disease 2019 (COVID-19), influenza A virus (InfA), syncytial B (RSVB), wastewater. Four cDNA input ratios (20 %-70 %) two extraction kits were evaluated optimization, with 30 % AllPrep PowerViral DNA/RNA Kit (Qiagen) exhibiting optimal performance. The cdPCR assay applied year-long wastewater surveillance study Japan (n = 52), revealing distinct trends prevalence viruses. NoV-GII detected 96 samples highest mean concentration (6.1 ± 0.6 log10 copies/L), while SARS-CoV-2 InfA 60 50 samples, respectively, reflected circulation these pathogens within community. Notably, RSVB less frequently (25 %), line fewer cases reported during period. concentrations EnV showed significant positive correlations hand foot mouth herpangina cases, respectively. no observed RSV COVID-19, possibly due testing RSVA more prevalent also cluster outbreaks. These findings demonstrated utility detecting provided insights into community trends, representing an advancement WBE.

Язык: Английский

Optimization of a 6-plex Crystal Digital PCR® assay and its application to simultaneous surveillance of enteric and respiratory viruses in wastewater DOI Creative Commons
Bikash Malla, Sadhana Shrestha, Eiji Haramoto

и другие.

The Science of The Total Environment, Год журнала: 2025, Номер 970, С. 178939 - 178939

Опубликована: Март 1, 2025

Multiplex digital PCR (dPCR) approaches are commonly employed in wastewater-based epidemiology (WBE) studies. However, optimizing the dPCR workflow is a critical step to ensure its reliability and accuracy before application. In this study, 6-plex Crystal Digital PCR® (cdPCR) was optimized for simultaneous detection of six epidemiologically important pathogens, including three enteric viruses, noroviruses genogroups I II (NoV-GI GII) enteroviruses (EnV), respiratory severe acute syndrome coronavirus 2 (SARS-CoV-2), which causative agent disease 2019 (COVID-19), influenza A virus (InfA), syncytial B (RSVB), wastewater. Four cDNA input ratios (20 %-70 %) two extraction kits were evaluated optimization, with 30 % AllPrep PowerViral DNA/RNA Kit (Qiagen) exhibiting optimal performance. The cdPCR assay applied year-long wastewater surveillance study Japan (n = 52), revealing distinct trends prevalence viruses. NoV-GII detected 96 samples highest mean concentration (6.1 ± 0.6 log10 copies/L), while SARS-CoV-2 InfA 60 50 samples, respectively, reflected circulation these pathogens within community. Notably, RSVB less frequently (25 %), line fewer cases reported during period. concentrations EnV showed significant positive correlations hand foot mouth herpangina cases, respectively. no observed RSV COVID-19, possibly due testing RSVA more prevalent also cluster outbreaks. These findings demonstrated utility detecting provided insights into community trends, representing an advancement WBE.

Язык: Английский

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