Ultracentrifugation as a direct method to concentrate viruses in environmental waters: virus-like particle enumeration as a new approach to determine the efficiency of recovery

Abstract

Some health important enteric viruses are considered to be emerging waterborne pathogens and so the improvement of detection of these viruses in the aquatic environment is one of the most important steps in dealing with these pathogens. Since these viruses may be present in low numbers in water, it is necessary to concentrate water samples before viral detection. Although there are several methods to concentrate viruses in environmental waters, all present some drawbacks and consequently the method should be chosen that, despite its limitations, is adequate to achieve the aim of each study. As the effectiveness of the concentration methods is evaluated by determining the efficiency of viral recovery after concentration, it is important to use a simple and effective approach to evaluate their recovery efficiency. In this work ultracentrifugation, usually used as a secondary step for virus concentration, was evaluated as the main method to concentrate directly viruses in environmental water samples, using the microscopic enumeration of virus-like particles (VLP) as a new approach to estimate the efficiency of recovery. As the flocculation method is currently employed to concentrate viruses in environmental waters, it was also used in this study to assess the efficiency of the ultracentrifugation as the main viral concentration method in environmental waters. The results of this study indicate that ultracentrifugation is an adequate approach to concentrate viruses directly from environmental waters (recovery percentages between 66 and 72% in wastewaters and between 66 and 76% in recreational waters) and that the determination of VLP by epifluorescence microscopy is a simple, fast and cheap alternative approach to determine the recovery efficiency of the viral concentration methods.

Routine screening of harmful microorganisms in beach sands: Implications to public health

“…Concentration of recreational water samples is performed by filtration through nitrocellulose membranes, elution, and organic flocculation (Wyn-Jones and Sellwood, 2001). Ultracentrifugation is also an efficient alternative concentration method (Prata et al, 2012). Another alternative method of filtration is through VIRADEL cartridges filter (Ortega et al, 2009), based on standard protocols (US EPA, 2001).…”

“…The concentration step is critical, since viruses may be present in low numbers in water (Prata et al, 2012). Concentration of recreational water samples is performed by filtration through nitrocellulose membranes, elution, and organic flocculation (Wyn-Jones and Sellwood, 2001).…”

A comprehensive method for amplicon-based and metagenomic characterization of viruses, bacteria, and eukaryotes in freshwater samples

“…Comparable recovery efficiencies have been reported with ultracentrifugation [68, 69]. Additional file 1: Figure S1 depicts quantitation of adenovirus (A) and enterovirus (B) per milliliter sample throughout different stages of the ultracentrifugation process and over time (1, 2, and 4 h).…”

Enteric Viruses as Contaminants and Bioindicators in Environmental Samples: A Review

“…However, Prata et al (2012) recently reported that the ultracentrifugation associated with the microscopic enumeration of virus-like particles (VLP) was an adequate approach to concentrate viruses directly from environmental waters (recovery percentages between 66 and 72% in wastewaters and between 66 and 76% in recreational waters).…”