Abstract

Standardization and validation of alternative cell lysis methods used for quantifying total cyanotoxins is needed to improve laboratory response time goals for total cyanotoxin analysis. In this study, five cell lysis methods (i.e., probe sonication, microwave, freeze-thaw, chemical lysis with Abraxis QuikLyseTM, and chemical lysis with copper sulfate) were assessed using laboratory-cultured Microcystis aeruginosa (M. aeruginosa) cells. Methods were evaluated for destruction of cells (as determined by optical density of the sample) and recovery of total microcystin-LR (MC-LR) using three M. aeruginosa cell densities (i.e., 1 × 105 cells/mL (low-density), 1 × 106 cells/mL (medium-density), and 1 × 107 cells/mL (high-density)). Of the physical lysis methods, both freeze-thaw (1 to 5 cycles) and pulsed probe sonication (2 to 10 min) resulted in >80% destruction of cells and consistent (>80%) release and recovery of intracellular MC-LR. Microwave (3 to 5 min) did not demonstrate the same decrease in optical density (<50%), although it provided effective release and recovery of >80% intracellular MC-LR. Abraxis QuikLyseTM was similarly effective for intracellular MC-LR recovery across the different M. aeruginosa cell densities. Copper sulfate (up to 500 mg/L Cu2+) did not lyse cells nor release intracellular MC-LR within 20 min. None of the methods appeared to cause degradation of MC-LR. Probe sonication, microwave, and Abraxis QuikLyseTM served as rapid lysis methods (within minutes) with varying associated costs, while freeze-thaw provided a viable, low-cost alternative if time permits.

Highlights

  • optical density at 730 nm (OD730) indicated that most cells were destroyed by 2 min (Figure 1b)

  • Cross-laboratory validation of sonication demonstrated that most cells were destroyed, and the majority of MC-LR was extracellular by 2 min (Figure S3)

  • Did not demonstrate the same decrease in OD730 correlated with cell destruction in this study, these methods resulted in effective release of intracellular MC-LR

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Summary

Introduction

Cyanotoxins, produced by cyanobacteria, present a risk to drinking water as recognized by health advisory levels established in the United States (U.S.) and several other countries [1,2,3]. Monitoring plans may include total, intracellular, and extracellular cyanotoxin measurements to assess concentrations already dissolved in the water supply (extracellular) and how much additional risk may be experienced following cell death (total/intracellular). Adsorption processes have been shown to cause shearing of cells, potentially resulting in the release of intracellular cyanotoxins [11]. Due to potential for release of intracellular cyanotoxins in either the source water or the treatment process, drinking-water utilities must consider total cyanotoxin concentrations for appropriate planning and risk management

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