Analysis of changes over time in microcystin content of nitrogen-limited Microcystis aeruginosa (Kützing) Lemmermann batch cultures (strain MASHOl-A19) showed that net microcystin production was limited to the phase of growth when cell concentration was increasing. The net microcystin production rate decreased as the specific cell division rate (µc) decreased, but, more importantly, the specific toxin production rate (µMCYST) decreased at an identical rate to that of µc when the culture became nitrate-limited. The actual size of the microcystin pool (total culture microcystin concentration) increased while cells were dividing, then remained constant or decreased only slightly during the stationary and death phases, even when the cultures were severely nitrate-starved. These findings demonstrate conclusively that the processes of cell division and microcystin production are tightly coupled under nitrogen-limited cell division. Our findings suggest that microcystin production is controlled by environmental effects on the rate of cell division, not through any direct effect on the metabolic pathways of toxin production. Reevaluation of data presented by others shows this to be the case for two other cyanobacterial species producing nine different microcystins over a wide range of environmental variables. We believe these relationships now provide a unifying view of environmental control of microcystin production in hepatotoxic cyanobacteria. We conclude that there is a direct linear correlation between cell division and microcystin production rates in all microcystin-producing cyanobacteria regardless of the environmental factor that is limiting cell division. We also conclude that microcystin is not a secondary metabolite, as is currently thought, but that it displays many of the attributes of essential intracellular nitrogenous compounds in toxigenic cyanobacteria.Microcystis aeruginosa (Kützing) Lemmermann is one of a number of species of cyanobacteria that may produce a suite of cyclic peptide hepatotoxins known as microcystins (Botes et al. 1984; Carmichael 1994). Because of growing public health concerns, there has been intense interest by scientists and water managers in elucidating the factors controlling the toxicity of blooms of M. aeruginosa and other cyanobacterial species (Carmichael 1992a(Carmichael , 1994.The gravimetric microcystin (MCYST) concentration of dried M. aeruginosa may vary by more than three orders of magnitude between individual bloom and culture isolates (Belch et al. 1997). Indeed, many isolates and blooms are reported to be nontoxic, reputedly containing no microcystin. The problem facing researchers is to ascertain whether the observed variation in microcystin content of field samples results from changes in the dominance of strains of varying microcystin content or from the influence of changing environmental variables on microcystin synthesis and metabolism. The literature provides evidence to support both mechanisms in culture and natural waters. Genetic variation is mos...

Internal pressurization and convective through-flow are demonstrated to be common attributes of wetland plants with cylindrical culms or linear leaves. Eight of 14 species tested produced static internal gas pressure differentials of 200-1,300 Pa relative to ambient and internal convective airflows of 0.2 to > 10 cm3 min-' culm-I, depending on species. Four species produced internal static pressure differentials of < 100 Pa. Two species did not pressurize. The driving forces are gradients in temperature and water vapor between the internal gas spaces of the plants and the ambient atmosphere (thermal transpiration and humidity-induced pressurization). A clear diel variation in pressurization and convective flow was observed; rates were highest in the afternoon and lowest at night, responding to ambient changes in light, temperature, and humidity. The resistance to airflow at the stem-rhizome junction was very high for some species, resulting in a low ability to convert internal pressurization into convective airflow through the rhizomes. Species with a high potential for internal pressurization and a low internal resistance to convective flow seem to have a competitive advantage over species that rely exclusively on diffisive gas transport, which allows them to grow in deeper waters.

Cell quotas of microcystin (Q MCYST ; femtomoles of MCYST per cell), protein, and chlorophyll a (Chl a), cell dry weight, and cell volume were measured over a range of growth rates in N-limited chemostat cultures of the toxic cyanobacterium Microcystis aeruginosa MASH 01-A19. There was a positive linear relationship between Q MCYST and specific growth rate (), from which we propose a generalized model that enables Q MCYST at any nutrient-limited growth rate to be predicted based on a single batch culture experiment. The model predicts Q MCYST from , max (maximum specific growth rate), Q MCYSTmax (maximum cell quota), and Q MCYSTmin (minimum cell quota). Under the conditions examined in this study, we predict a Q MCYSTmax of 0.129 fmol cell ؊1 at max and a Q MCYSTmin of 0.050 fmol cell ؊1 at ‫؍‬ 0. Net MCYST production rate (R MCYST ) asymptotes to zero at ‫؍‬ 0 and reaches a maximum of 0.155 fmol cell The microcystins (MCYSTs) are a group of cyclic heptapeptide toxins produced by several cyanobacterial species. Of the more than 60 MCYSTs characterized to date (19,27,29), most are potent inhibitors of protein phosphatases 1 and 2A from both plants and animals (17). One of the most common MCYST-producing cyanobacteria is the bloom-forming Microcystis aeruginosa (Kützing) Lemmermann. Due to the widespread distribution and potential toxicity of this species (toxic strains have been found worldwide), M. aeruginosa has been implicated in a number of animal-poisoning incidents (e.g., reference 7) and more recently in human fatalities (11,23).M. aeruginosa is a unicellular, colonial freshwater cyanobacterium which often forms blooms during warmer months in eutrophic lakes and reservoirs (37). For this reason, much research has been concerned with the environmental factors which lead to bloom formation and toxin production in this species. A wide range of batch culture studies have shown that the variables influencing MCYST content include trace metal supply (15), nitrogen (N) and phosphorus (P) (31), light and temperature (38), and pH (34). Comparative studies on MCYST production by M. aeruginosa in continuous culture, however, have been limited to examination of the effects of photon irradiance (35), N, P, and Fe 3ϩ limitation (16, 36), and more recently P limitation (20). Despite this considerable pool of data concerning MCYST production, few studies (with the exception of the work carried out by Rapala and coworkers [25,26]) have been able to quantitatively relate MCYST content to any growth determinant.In a previous batch culture study, we presented data on the effect of N supply on the cellular production of MCYSTs (21). This work showed that the net specific rate of MCYST production was equal to the cell specific growth rate. The application of these findings to previously published batch culture studies suggested that the relationship held under a variety of culture conditions and that MCYST production was indirectly affected by environmental factors through their effects on cell division. A consequence of this l...

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Morphological, toxicological, and genetic variation was examined among 19 strains of Nodularia. The strains examined could be morphologically discriminated into four groups corresponding to N. spumigena Mertens, N. sphaerocarpa Bornet et Flahault, and two strains that did not clearly correspond to currently accepted Nodularia species. Genetic variation was examined using nucleotide sequencing of the phycocyanin intergenic spacer region (cpcBA‐IGS) and RAPD‐PCR. The PCR‐RFLP of the cpcBA‐IGS differentiated four genotypes corresponding to the four morphological groups. However, nucleotide sequencing of 598 bp of the 690‐bp fragment showed that one of the three strains corresponding to N. sphaerocarpa (PCC 7804) was genetically divergent from the other two, suggesting that it constitutes a distinct species. Nucleotide variation within the morphospecies groups was limited (<1%), and all 14 Australian strains of N. spumigena possessed identical cpcBA‐IGS sequences. The RAPD‐PCR differentiated the same groups as the cpcBA sequencing and discriminated each of the seven different Australian populations of N. spumigena. Strains from within a bloom appeared genetically identical; however, strains isolated from different blooms could be separated into either a western or a southeastern Australian cluster, with one strain from western Australia showing considerable genetic divergence. The pattern of variation suggests that individual blooms of N. spumigena are clonal but also that Australian N. spumigena populations are genetically distinct from each other. Examination of genetic distance within and between blooms and within and between morphological groups showed clear genetic dicontinuities that, in combination with the cpcBA‐IGS data, suggest that Nodularia contains genetically distinct morphospecies rather than a continuous cline of genetic variation. Furthermore, these morphospecies are genetically variable, exhibiting hierarchical patterns of genetic variation on regional and global scales. Production of the hepatotoxin nodularin was not restricted to one genetic lineage but was distributed across three of the five genotypic groups. A strain of N. spumigena from a nontoxic Australian population was found to fall within the range of genetic variation for other toxic Australian strains and appears to be a unique nontoxic strain that might have arisen by loss of toxin production capacity.

A mixed bloom of Microcystis aeruginosa forma aeruginosa and forma flos-aquae from the Swan River, Western Australia, was confirmed toxic by HPLC analysis. At least four, and possibly 11, microcystins were detected in cell-free extracts. Live bloom material was cultured at salt concentrations up to 21.2 g L–1 (total salts). The cultures were salt tolerant up to 9.8 g L–1. Reduction in the total cell concentration in the first 23 h was only observed in the highest salt treatment and first-order rate constants for cell lysis were higher than the rates for reduction of the intracellular microcystin pool size for that treatment. This suggests preferential lysis of genotypes with lower salinity tolerance and toxigenicity. This increased the toxicity of the mixed bloom population and the apparent microcystin cell quota without any change to the intracellular microcystin pool size. Therefore, the toxicity of bloom material may change through preferential lysis of cells with lower tolerances to changing environmental conditions, including salinity. Managers should be aware that the World Health Organization alert levels of 105 cells mL–1 for human contact exposure to cyanobacteria may not be a suitable prima facie test during these periods.

The worldwide appearance of toxic cyanobacterial blooms in drinking water supplies has raised concerns about systemic effects on human health. Conventional water treatment methods are poor at removing low concentrations of cyanotoxins, and specialized treatment is usually necessary for treatment of contaminated water. In this study, the applicability of heterogeneous photocatalytic degradation of low concentrations of the cyantoxin microcystin-LR in a natural organic-aqueous matrix is examined using titanium dioxide as the photocatalyst. The initial rate of toxin degradation is strongly pH dependent in a manner mirrored by the pH dependence of toxin adsorption to TiO 2 . Rapid degradation of toxin occurs in the acidic pH range in the presence of light and TiO 2 with a maximum initial rate of degradation occurring at pH 3.5, while at higher pH, a distinct lag is observed prior to commencement of toxin degradation. It is proposed that in the pH range where microcystin-LR adsorbs to TiO 2 , it is degraded principally by long-lived organic radicals generated through oxidation of adsorbed cyanobacterial exudate. At higher pH, where microcystin-LR adsorption to TiO 2 is insignificant, it is proposed that these organic radicals diffuse into solution and (after a lag) initiate oxidation of the toxin in dissolved phase.