Fluorescence instrument for in situ monitoring of viral abundance in water, wastewater and recycled water

In a world of advanced molecular methods quantifying viruses in water remains one of the most inefficient and costly. Using a general molecular DNA/RNA probe – SYBR Gold combined with differential filtration a fast, cost effective and sensitive method is presented to determine the concentration of viruses in water in situ or on-line. The approach differentiates the nucleotide size fractions that are stained with SYBR Gold to show only those associated with Viral DNA and RNA. There was a linear relationship between the fluorescence maxima for SYBR Gold added to wastewater and viral numbers determined with direct counting using epifluorescent microscopy (r2 = 0.97) and for a range of diverse natural water samples (r2 = 0.86). The method was applied to water from the tropics and Antarctica, marine and freshwater environments where natural viral abundances ranged from 106 to 108 viruses mL−1. The method takes into account the background fluorescence that represented 70% of total fluorescence and any auto-fluorescence due to other dissolved organic carbon. While DNAse II lowered the background fluorescence associated with free DNA and RNA it could not be eliminated. The technique presented is suitable for monitoring in situ viral numbers in natural water bodies and engineered water treatment processes. This on-line viral monitoring design has the potential to replace human viral pathogen indicators.

► I describe aquatic viral DNA/RNA size fractions labeled with a SYBR fluorescence dye. ► Viral numbers correlate to fluorescence if background (<10 nm size fraction) removed. ► Present an accurate and sensitive measure of viral abundance in situ. ► Show on-line fluorescence method to quantify viruses in engineered and natural waters. ► Describe portable, inexpensive fluorescence instrument to quantifying aquatic viruses.