Diversity of cyanobacteria and cyanotoxins in Hartbeespoort Dam, South Africa

Abstract

Abstract. The South African Hartbeespoort Dam is known for the occurrence of heavy Microcystis blooms. Although a few other cyanobacterial genera have been described, no detailed study on those cyanobacteria and their potential toxin production has been conducted. The diversity of cyanobacterial species and toxins is most probably underestimated. To ascertain the cyanobacterial composition and presence of cyanobacterial toxins in Hartbeespoort Dam, water samples were collected in April 2011. In a polyphasic approach, 27 isolated cyanobacterial strains were classified morphologically and phylogenetically and tested for microcystins (MCs), cylindrospermopsin (CYN), saxitoxins (STXs) and anatoxin-a (ATX) by liquid chromatography-tandem mass spectrometry (LC-MS/MS) and screened for toxin-encoding gene fragments. The isolated strains were identified as Sphaerospermopsis reniformis, Sphaerospermopsis aphanizomenoides, Cylindrospermopsis curvispora, Raphidiopsis curvata, Raphidiopsis mediterrranea and Microcystis aeruginosa. Only one of the Microcystis strains (AB2011/53) produced microcystins (35 variants). Forty-one microcystin variants were detected in the environmental sample from Hartbeespoort Dam, suggesting the existence of other microcystin producing strains in Hartbeespoort Dam. All investigated strains tested negative for CYN, STXs and ATX and their encoding genes. The mcyE gene of the microcystin gene cluster was found in the microcystin-producing Microcystis strain AB2011/53 and in eight non-microcystin-producing Microcystis strains, indicating that mcyE is not a good surrogate for microcystin production in environmental samples.

Variability in the sxt Gene Clusters of PSP Toxin Producing Aphanizomenon gracile Strains from Norway, Spain, Germany and North America

“…A modified isolation of genomic DNA was conducted after Ballot et al [32]. Instead of horizontal vortexing, a bead beating step (3 × 30 sec, 6700 rpm) in a Precellys 24 Beadbeater (Bertin, Technologies, Saint Quentin, France), was used to disrupt the cells.…”

Anatoxin-a producing Tychonema (Cyanobacteria) in European waterbodies

“…These primers proved to be useful also in the amplification of the anaC genes in Tychonema, suggesting the potential for a wider application of these specific protocols also to other Oscillatoriales. Similar considerations apply to the atxoaf r primers, which were originally designed to detect Aphanizomenon species and other cyanobacteria (Ballot et al, 2010(Ballot et al, , 2014. In perspective, the comparison of the ana gene cluster encoding ATX in Tychonema and in the other cyano bacteria will provide further insight for the design of protocols tailored for the detection of pelagic ATX producers (cf.…”

“…strains were investigated for the production of MC and ATX using the Abraxis Microcystins/Nodularins (ADDA), ELISA Kit and the Abraxis Anatoxin a Receptor Binding Assay (Biosense, Bergen, Norway) respectively. The same strains were tested for the anaF encoding gene using the primers atxoaf and atxoar and the protocol according to Ballot et al (2010Ballot et al ( , 2014. The presence of mcyE genes was evaluated according to the PCR protocols of Rantala et al (2006) using general primers (mcyE F2/R4).…”

“…The presence of mcyE genes was evaluated according to the PCR protocols of Rantala et al (2006) using general primers (mcyE F2/R4) and Anabaena 90 as a positive control (Ana 90; UHCC). The presence of anatoxin a synthe tase gene (anaC) was determined using the primer pairs anaC osc, Osc 193 {UHCC) as positive control, and the methods described in Rantala Ylinen et al (2011b Ballot et al (2014), PCR and sequencing of the 165 rRNA gene was conducted using the methods described in Ballot et al (2008). The Norwegian Tychonema sp.…”

Sulfide Oxidations for LC-MS Analysis of Methionine-Containing Microcystins in Dolichospermum flos-aquae NIVA-CYA 656

“…14 Recently, we described a method using thiol derivatization together with LC-MS/MS analysis to identify microcystins in complex matrices, based on the reactivity of the Mdha and Dha groups ( Figure 1) present in most MCs. 12,16,17 This method also discriminates between microcystins containing the thiol-reactive Mdha and Dha and the unreactive Mdhb and Dhb (Figure 1) residues, which are otherwise difficult to differentiate by LC-MS. 18 Microcystins containing methionine are relatively uncommon in the literature, but MC-M(O)R (7), 19 While analyzing an extract from D. f los-aquae by LC-MS 2 , we observed peaks at m/z 1015.5, 1029.5, 999.5, and 1013.5 whose thiol reactivities (Table 1, Supporting Information Figure S41) and MS 2 spectra (Supporting Information Figures S25 and S27−29) were consistent with Met-containing microcystins, but in the absence of high-resolution MS, these masses also corresponded to plausible non-Met-containing microcystins.…”