An inexpensive, accurate, and precise wet-mount method for enumerating aquatic viruses

Area of Science:

• Environmental microbiology
• Aquatic virology

Background:

• Viruses play crucial roles in microbial ecology, influencing biogeochemical cycles and microbial populations.
• Accurate quantification of viral abundance is essential for understanding these ecological impacts.
• Current methods like the filter mount technique are effective but costly.

Purpose of the Study:

• To develop and validate a cost-effective alternative method for enumerating viruses in aquatic environments.
• To compare the accuracy and precision of the new method against the established filter mount technique.

Main Methods:

• A novel wet-mount method involving direct slide mounting of fluorescently stained samples.
• Optional chemical flocculation for samples with lower viral concentrations (<5×10(7) viruses ml(-1)).
• Enumeration using epifluorescence microscopy and quantification via a ratio of viruses to known concentration microsphere beads.

Main Results:

• The wet-mount method demonstrated significant correlation and equivalent precision to the filter mount method.
• The new method is approximately 500-fold less expensive in materials cost.
• Accurate virus enumeration was achieved across a wide range of concentrations (2.17×10(6) to 1.37×10(8) viruses ml(-1)) in both marine and freshwater samples.

Conclusions:

• The wet-mount method provides a rapid, precise, and accurate alternative for aquatic virus enumeration.
• This technique is suitable for field and laboratory applications, especially for samples with viral concentrations ≥1×10(6) viruses ml(-1).
• The significantly lower cost makes viral quantification more accessible for ecological studies.