Immuno-gold localization of hepatotoxins in cyanobacterial cells

Abstract

A polyclonal antibody against the potent hepatotoxic cyclic peptide microcystins and nodularins was used in conjunction with immuno-gold labelling to localize the toxins in three strains of cyanobacteria. Ultrastructurally, there were no major differences between unicellular Microcystis aeruginosa strain PCC 7820 (toxin-producing strain) and M. aeruginosa strain UTEX 2063 (non-toxin-producing strain), except that M. aeruginosa PCC 7820 cells had a sheath. The thickness of the sheath was about 12 nm and was distinguishable from the cell wall at the ultrastructural level only when the specimen was stained en bloc with uranyl acetate. Microcystins and nodularin were found in M. aeruginosa PCC 7820 and Nodularia spumigena strain L-575 respectively, but not in nontoxic M. aeroginosa UTEX 2063. In M. aeruginosa PCC 7820 cells, microcystin was found primarily in the thylakoid area and nucleoid, with smaller amounts in the cell wall and sheath. Only nonspecific labelling was found in other cellular inclusions, such as polyhedral bodies, cyanophycin granules and membrane-limited inclusions. In strain N. spumigena L-575, nodularin was found in both vegetative cells and heterocysts with a distribution similar to that in M. aeruginosa PCC 7820.

Effects of nonylphenol on the growth and microcystin production of Microcystis strains

“…On the other hand, low concentration of NP (0.05-0.2 mg/L) stimulated slightly to the synthesis of chlorophyll a and carotinoid of toxic strain PCC7820. Shi et al (1995) found that microcystin was associated with the thylakoid membranes of M. aeruginosa PCC7820, suggesting a close physiological association between microcystins and the photosynthetic machinery of the cell. The finding in our study may suggest that NP had taken part in the physiological activity of some toxic M. aeruginosa cells and such activity had a certain extent relationship with toxin production.…”

“…The reason is not clear. The observation by Shi et al (1995) of some localization of microcystin in the cell wall and sheath suggest that microcystin may act as transmembrane transporters of calcium, magnesium or other metals. Stimulation of membrane receptors is believed to be a cause for the activation of cell functions.…”

Quantification and localization of microcystins in colonies of a laboratory strain ofMicrocystis(Cyanobacteria) using immunological methods

“…Recent MC-localization studies have used cryosectioning to prepare cells for immunogold labelling and electron microscopy since this technique avoids using organic solvents, which extract MC from cells . Shi et al (1995) and Young et al (2005) found that most of the intracellular pool of MCs in Microcystis, as determined by immunogold label, is localized in the thylakoid region, possibly indicating a role for MCs in the assembly, structure or function of the photosynthetic apparatus, or in other light-related processes. High labelling densities were also found around polyphosphate bodies .…”

“…Antisera raised against MC-LR have been successfully used in immunoelectron microscopy to localize MCs in laboratory cultures of Microcystis spp. (Shi et al, 1995;Young et al, 2005;Gerbersdorf, 2006). Traditional preparative methods for immunoelectron microscopy (e.g.…”

“…Traditional preparative methods for immunoelectron microscopy (e.g. Shi et al, 1995) extract MCs from fixed samples during the ethanol dehydration step . An alternative means of preparing cells for electron microscopy, cryosectioning, is increasingly being used and is considered to be the most useful technique for immuno-localization studies with respect to membrane visibility and labelling sensitivity (Koster & Klumperman, 2003).…”