A high-throughput method for counting cyanobacteria using fluorescence microscopy

Abstract

Harmful algal blooms (HABs) occur all over the world naturally, but are increasing in frequency largely due to eutrophication. The U.S. Army Corps of Engineers (USACE) manages over 400 freshwater waterways making cyanobacteria HABs (cyanoHABs) a significant threat to water quality. CyanoHABs have been identified as a significant problem in USACE managed waterways where districts have reported associated fish kills and waterway closures. For USACE districts, when contracted out, the return time for cyanobacterial enumeration is approximately 30 days, therefore, there is a need to provide a better approach to counting cyanobacteria. However, counting cyanobacteria is notoriously difficult due to large filaments that make conventional counting methods impractical. It is anticipated that the application of this method for routine counting of field samples will greatly reduce analysis time, and therefore, will enhance the ability to combat HABs. In addition to cyanoHAB monitoring, cell densities are required for various down-stream techniques, including nucleic acid extractions where overloading of solid phase extraction columns can cause a loss of sample. This study developed automated image processing routines that enable rapid and accurate cell counting. The results of this method compared favorably with manual counting of single-cell and filamentous cyanobacteria cultures at a fraction of the analysis time. DISCLAIMER: The contents of this report are not to be used for advertising, publication, or promotional purposes. Citation of trade names does not constitute an official endorsement or approval of the use of such commercial products. All product names and trademarks cited are the property of their respective owners. The findings of this report are not to be construed as an official Department of the Army position unless so designated by other authorized documents. DESTROY THIS REPORT WHEN NO LONGER NEEDED. DO NOT RETURN IT TO THE ORIGINATOR.