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Title: Capsid integrity qPCR—an Azo-dye based and culture-independent approach to estimate adenovirus infectivity after disinfection and in the aquatic environment
Authors: Leifels, Mats
Shoults, David
Wiedemeyer, Alyssa
Ashbolt, Nicholas J.
Sozzi, Emanuele
Hagemeier, Angela
Jurzik, Lars
Keywords: Capsid Integrity qPCR
Human Adenovirus
Issue Date: 2019
Source: Leifels, M., Shoults, D., Wiedemeyer, A., Ashbolt, N. J., Sozzi, E., Hagemeier, A., & Jurzik, L. (2019). Capsid Integrity qPCR—An Azo-Dye Based and Culture-Independent Approach to Estimate Adenovirus Infectivity after Disinfection and in the Aquatic Environment. Water, 11(6), 1196-. doi:10.3390/w11061196
Series/Report no.: Water
Abstract: Recreational, reclaimed and drinking source waters worldwide are under increasing anthropogenic pressure, and often contain waterborne enteric bacterial, protozoan, and viral pathogens originating from non-point source fecal contamination. Recently, the capsid integrity (ci)-qPCR, utilizing the azo-dyes propidium monoazide (PMA) or ethidium monoazide (EMA), has been shown to reduce false-positive signals under laboratory conditions as well as in food safety applications, thus improving the qPCR estimation of virions of public health significance. The compatibility of two widely used human adenovirus (HAdV) qPCR protocols was evaluated with the addition of a PMA/EMA pretreatment using a range of spiked and environmental samples. Stock suspensions of HAdV were inactivated using heat, UV, and chlorine before being quantified by cell culture, qPCR, and ci-qPCR. Apparent inactivation of virions was detected for heat and chlorine treated HAdV while there was no significant difference between ci-qPCR and qPCR protocols after disinfection by UV. In a follow-up comparative analysis under more complex matrix conditions, 51 surface and 24 wastewater samples pre/post UV treatment were assessed for enteric waterborne HAdV to evaluate the ability of ci-qPCR to reduce the number of false-positive results when compared to conventional qPCR and cell culture. Azo-dye pretreatment of non-UV inactivated samples was shown to improve the ability of molecular HAdV quantification by reducing signals from virions with an accessible genome, thereby increasing the relevance of qPCR results for public health purposes, particularly suited to resource-limited low and middle-income settings.
ISSN: 2073-4441
DOI: 10.3390/w11061196
Rights: © 2019 by the Authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (
Fulltext Permission: open
Fulltext Availability: With Fulltext
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