An On-Farm Workflow for Predictive Management of Paralytic Shellfish Toxin-Producing Harmful Algal Blooms for the Aquaculture Industry

Abstract

Paralytic shellfish toxins (PSTs) produced by marine dinoflagellates significantly impact shellfish industries worldwide. Early detection on-farm and with minimal training would allow additional time for management decisions to minimize economic losses. Here, we describe and test a standardized workflow based on the detection of sxtA4, an initial gene in the biosynthesis of PSTs. The workflow is simple and inexpensive and does not require a specialized laboratory. It consists of (1) water collection and filtration using a custom gravity sampler, (2) buffer selection for sample preservation and cell lysis for DNA, and (3) an assay based on a region of sxtA, DinoDtec lyophilized quantitative polymerase chain reaction (qPCR) assay. Water samples spiked with Alexandrium catenella showed a cell recovery of >90% when compared to light microscopy counts. The performance of the lysis method (90.3% efficient), Longmire's buffer, and the DinoDtec qPCR assay (tested across a range of Alexandrium species (90.7-106.9% efficiency; r² > 0.99)) was found to be specific, sensitive, and efficient. We tested the application of this workflow weekly from May 2016 to 30th October 2017 to compare the relationship between sxtA4 copies L^-1 in seawater and PSTs in mussel tissue (Mytilus galloprovincialis) on-farm and spatially (across multiple sites), effectively demonstrating an ∼2 week early warning of two A. catenella HABs (r = 0.95). Our tool provides an early, accurate, and efficient method for the identification of PST risk in shellfish aquaculture.

Keywords: alexandrium spp.; aquaculture industry; harmful algal blooms (HABs); molecular detection; on-farm workflow; paralytic shellfish toxins (PSTs); sxtA4 gene.

Figure 1

Photo and technical diagram of the gravity-operated portable water sampler showing the following components: A. a first-stage prefilter with a 100 μm mesh; B. a 3 liter water chamber; and C. a second-stage filter with an 11 μm mesh designed to capture and retain HAB cells such as dinoflagellates and all measurements. The sampler is made from PVC piping with a midline ball valve/tap.

Figure 2

Standard curve of the DinoDtec assay using lysed cells of A. pacificum strain MMWA 83 as the template DNA, showing the quantification cycle (y-axis) versus the known cell number in log-scale (x-axis).

Figure 3

Copy number of sxtA4 L^–1 in seawater and total concentration of PST in mussel tissue (mg kg^–1) during the 2016 A. catenella bloom from Spring Bay Seafoods. The inset shows a portion of the same data from June to July 2016, with sxtA4 L^–1 in seawater on a log-scale, highlighting that the qPCR assay detected significant sxtA4 copies prior to the detection of toxins in mussel tissue.

Figure 4

Dynamics of sxtA4 L^–1 copies in seawater and the total concentration of PST in mussel tissue during the 2017 A. catenella bloom from Spring Bay Seafoods.

Figure 5

Sampling stations of the RV Southern Cross survey along the east coast of Tasmania, Australia, with the concentration of surface-sxtA4 copy L^–1 detected at each site during the peak (26–28 August) of the A. catenella bloom in 2016.