Monitoring changing toxigenicity of a cyanobacterial bloom by molecular methods

Abstract

Cyanobacterial blooms are potential health hazards in water supply reservoirs. This paper reports analyses of a cyanobacterial bloom by use of PCR-based methods for direct detection and identification of strains present and determination of their toxigenicity. Serial samples from Malpas Dam, in the New England region of Australia, were analyzed during a prolonged, mixed cyanobacterial bloom in the summer of 2000 to 2001. Malpas Dam has been shown in the past to have toxic blooms of Microcystis aeruginosa that have caused liver damage in the human population drinking from this water supply reservoir. Cyanobacterial genera were detected at low cell numbers by PCR amplification of the phycocyanin intergenic spacer region between the genes for the beta and alpha subunits. The potential for microcystin production was determined by PCR amplification of a gene in the microcystin biosynthesis pathway. The potential for saxitoxin production was determined by PCR amplification of a region of the 16S rRNA gene of Anabaena circinalis strains. Toxicity of samples was established by mouse bioassay and high-pressure liquid chromatography. We show that bloom components can be identified and monitored for toxigenicity by PCR more effectively than by other methods such as microscopy and mouse bioassay. We also show that toxigenic strains of Anabaena and Microcystis spp. occur at this site and that, over the course of the bloom, the cell types and toxicity changed. This work demonstrates that PCR detection of potential toxicity can enhance the management of a significant public health hazard.

FIG. 1.

Cyanobacterial cell counts for site 1 surface samples, summer 2000 to 2001. Arrows indicate sampling dates shown in Table 2. Data are summarized from ADC records.

FIG. 2.

Identification of bloom samples by PC-IGS sequences. Key bloom samples MD-80, MD-88, and MD-108 were compared with a reference culture and two database entries (see Tables 1 and 2 for more information). Sample MD-80 was confirmed as A. circinalis, and samples MD-88 and MD-108 are Microcystis species.

FIG. 3.

Detection of the NMT domain of the microcystin synthetase gene, mcyA, in Malpas Dam bloom samples (Table 2) and control samples (Table 1). Of the control samples, only M. aeruginosa PCC 7806 (M.a. 7806) is a microcystin producer.

FIG. 4.

Detection of potentially saxitoxin producing A. circinalis strains in Malpas Dam bloom samples by amplification of a region of the 16S rRNA gene. For descriptions of strain and bloom samples, see Tables 1 and 2. Of the four reference strains of A. circinalis, AWQC118C and AWQC131C are known saxitoxin producers while AWQC271C and AWQC306A are not known to produce saxitoxin, although AWQC271C gives a positive result in this test (6).