Adenovirus and Norovirus in water and sediments of recreational aquatic environments: Comparison of concentration, recovery and detection methods

LUCÍA CHÁVEZ DÍAZ; MACHADO FUMIÁN, TULIO; POMA, HUGO RAMIRO; DOLORES GUTIÉRREZ CACCIABUE; MIAGOSTOVICH, MARIZE; RAJAL, VERONICA BEATRIZ

Florianópolis

Simposio; IV Simposio Latinoamericano de Virología Ambiental; 2018

Aquatic environments are intensively used for public recreational activities. In some cases, rivers and lakes become a risk to population due to the presence of some viruses that transmit waterborne diseases. Pathogens are not only found in water but also in sediments, which may act as a reservoir. An eventually resuspension of sediments could return microorganism to the water column. Since viruses are in general at low amounts in natural aquatic environments, a concentration step before the detection is necessary. The aim of this work was to analyze different methods to detect adenovirus and norovirus genogroup I and II in two recreational aquatic environments in Salta, Argentina. Water and sediment samples (3 replicates per occasion) were collected from December 2015 to September 2016 from the Wierna River (WR) (5 samples, 15 in total) and General Belgrano Reservoir (GBR) (6 samples, 18 in total). Water samples were concentrated by ultrafiltration using PP7 recovery control and nucleic acids were extracted with commercial QIAamp Viral RNA Mini Kit (Qiagen, Germany). Adenovirus was analyzed by real-time PCR (qPCR) with two different sets of previously published primers (here referred as OS1 and OS2). Norovirus GI and GII were analyzed with the same primers and probes, but in the first case a RT was carried out followed by qPCR, and in the second a one-step quantitative multiplex RT-PCR system was used. For sediments, two nucleic acids extraction methods were used: i) Mobio PowerSoil DNA kit; and ii) skimmed-milk flocculation method (using PP7 as a recovery control), followed by nucleic acidsextraction with QIAamp Viral RNA Mini Kit. Using OS1, which targeted only gastrointestinal-related species (HAdV-F types 40 and 41), 13% and 11% positives cases of adenovirus in water were found at WR and GBR, respectively. Instead using OS2, which targeted all human adenovirus species, 40% and 83% positives samples were detected in water at each place, respectively. Regarding norovirus detection, only GII was detected in 20% and 11% of water samples from WR and GBR, respectively. Using the skimmed-milk flocculation method in sediments, adenovirus was detected in all samples of both environments, and norovirus GII was detected in one river and one reservoir samples. Only one positive sample for adenovirus was found using the MoBio kit. The desorption-concentration step in sediments seemed to be crucial for the recovery and detection of viruses.