Strains Of Cyanobacteria Research Articles

Schizopeptin 791, a new anabeanopeptin-like cyclic peptide from the cyanobacterium Schizothrix sp.

Schizopeptin 791 (1), a new trypsin inhibitor, related in structure to the anabeanopeptins, was isolated from the hydrophilic extract of the cultured terrestrial cyanobacterium Schizothrix sp. Homo- and heteronuclear 2D NMR techniques as well as HRFABMS determined the gross structure of 1. The relative and absolute stereochemistry of 1 was deduced from a combination of spectral data and Marfey's method for HPLC. Prior to this finding, anabeanopeptins have been isolated only from water-bloom-forming strains of cyanobacteria.

A molecular test of cyanobacterial phylogeny: inferences from constraint analyses

Parsimony and neighbor-joining analyses of 16S rDNA nucleotide sequences of 68 species and strains of cyanobacteria and prochlorophytes supported a monophyletic Nostocales, a monophyletic Stigonematales, three independent lineages of prochlorophytes within the cyanobacteria, and a paraphyletic Chroococcales (p<0.0001) and Oscillatoriales (p = 0.0147). Within the Oscillatoriales, the genus Oscillatoria formed an unnatural taxon (p<0.0001) and needs major revisions. Using constraint analysis, the genus Microcystis was found to be monophyletic and the paraphyletic positions of Microcystis elabens and M. holsatica are probably due to long-branch attraction. Further, a separation of Chroococcales based on varying levels of polyunsaturated fatty acids is more consistent with nucleotide-based phylogenies than with existing morphological groupings. It is proposed that Chroococcales be redefined to exclude the genus Microcystis, and that a new order be erected for Microcystis. Finally, it is more parsimonious to assume a common cyanobacterial/prochlorophyte ancestor, than to evoke de novo synthesis of chlb in each prochlorophyte lineage plus in the lineage leading to green chloroplasts. This common ancestor is proposed to have contained both chlorophyll a and b plus phycobilins. Subsequent multiple losses of chlb in cyanobacteria and the loss of chla and phycobilins in prochlorophytes, led to the currently observed pigment distribution. It is therefore, recommended that Prochlorales be reclassified as cyanobacteria.

Preliminary studies of cyanobacteria, picoplankton, and virioplankton in the Salton Sea with special attention to phylogenetic diversity among eight strains of filamentous cyanobacteria

Cyanobacterial diversity in the Salton Sea, a high-salinity, eutrophic lake in Southern California, was investigated using a combination of molecular and morphological approaches. Representatives of a total of 10 described genera (Oscillatoria, Spirulina, Arthrospira, Geitlerinema, Lyngbya, Leptolyngbya, Calothrix, Rivularia, Synechococcus, Synechocystis) were identified in the samples; additionally, the morphology of two cultured strains do not conform to any genus recognized at present by the bacteriological system. Genetic analysis, based on partial 16S rRNA sequences suggested considerable cryptic genetic variability among filamentous strains of similar or identical morphology and showed members of the form-genus Geitlerinema to be distributed among three major phylogenetic clades of cyanobacteria. Cyanobacterial mats, previously described from the Sea were, in fact, composed of both filamentous cyanobacteria and a roughly equivalent biomass of the sulfur-oxidizing bacterium Beggiatoa, indicating their formation in sulfide rich regions of the lake. Flow cytometric analysis of the water samples showed three striking differences between samples from the Salton Sea and representative marine waters: (1) phycoerythrin-containing unicells, while abundant, were much less abundant in the Salton Sea than they were in typical continental shelf waters, (2) Prochlorococcus appears to be completely absent, and (3) small (3–5 μm) eukaryotic algae were more abundant in the Salton Sea than in typical neritic waters by one-to-two orders-of-magnitude. Based on flow cytometric analysis, heterotrophic bacteria were more than an order of magnitude more abundant in the Salton Sea than in seawater collected from continental shelf environments. Virus particles were more abundant in the Salton Sea than in typical neritic waters, but did not show increases proportionate with the increase in bacteria, picocyanobacteria, or eukaryotic algae.

Role of NtcB in activation of nitrate assimilation genes in the cyanobacterium Synechocystis sp. strain PCC 6803.

In Synechocystis sp. strain PCC 6803, the genes encoding the proteins involved in nitrate assimilation are organized into two transcription units, nrtABCD-narB and nirA, the expression of which was repressed by ammonium and induced by inhibition of ammonium assimilation, suggesting involvement of NtcA in the transcriptional regulation. Under inducing conditions, expression of the two transcription units was enhanced by nitrite, suggesting regulation by NtcB, the nitrite-responsive transcriptional enhancer we previously identified in Synechococcus sp. strain PCC 7942. The slr0395 gene, which encodes a protein 47% identical to Synechococcus NtcB, was identified as the Synechocystis ntcB gene, on the basis of the inability of an slr0395 mutant to rapidly accumulate the transcripts of the nitrate assimilation genes upon induction and to respond to nitrite. While Synechococcus NtcB strictly requires nitrite for its action, Synechocystis NtcB enhanced transcription significantly even in the absence of nitrite. Whereas the Synechococcus ntcB mutant expresses the nitrate assimilation genes to a significant level in an NtcA-dependent manner, the Synechocystis ntcB mutant showed only low-level expression of the nitrate assimilation genes, indicating that NtcA by itself cannot efficiently promote expression of these genes in Synechocystis. Activities of the nitrate assimilation enzymes in the Synechocystis ntcB mutant were consequently low, being 40 to 50% of the wild-type level, and the cells grew on nitrate at a rate approximately threefold lower than that of the wild-type strain. These results showed that the contribution of NtcB to the expression of nitrate assimilation capability varies considerably among different strains of cyanobacteria.

Membrane dynamics as seen by Fourier transform infrared spectroscopy in a cyanobacterium, Synechocystis PCC 6803: The effects of lipid unsaturation and the protein-to-lipid ratio

The roles of lipid unsaturation and lipid-protein interactions in maintaining the physiologically required membrane dynamics were investigated in a cyanobacterium strain, Synechocystis PCC 6803. The specific effects of lipid unsaturation on the membrane structure were addressed by the use of desaturase-deficient (desA−/desD−) mutant cells (which contain only oleic acid as unsaturated fatty acid species) of Synechocystis PCC 6803. The dynamic properties of the membranes were determined from the temperature dependence of the symmetric CH2 stretching vibration frequency, which is indicative of the lipid fatty acyl chain disorder. It was found that a similar membrane dynamics is maintained at any growth temperature, in both the wild-type and the mutant cell membranes, with the exception of mutant cells grown at the lower physiological temperature limit. It seems that in the physiological temperature range the desaturase system of the cells can modulate the level of lipid desaturation sufficiently to maintain similar membrane dynamics. Below the range of normal growth temperatures, however, the extent of lipid disorder was always higher in the thylakoid than in the cytoplasmic membranes prepared from the same cells. This difference was attributed to the considerable difference in protein-to-lipid ratio in the two kinds of membranes, as determined from the ratio of the intensities of the protein amide I band and the lipid ester C=O vibration. The contributions to the membrane dynamics of an ab ovo present ‘structural’ lipid disorder due to the protein–lipid interactions and of a thermally induced ‘dynamic’ lipid disorder could be distinguished.

Harmful cyanobacterial toxic blooms in waste stabilisation ponds

A coccoid picocyanobacterium Synechocystis sp. (0.6-2 μm of cell diameter) was found to be dominant during summer period in the experimental wastewater stabilisation pond of Marrakesh. The taxonomy of this isolated strain was confirmed by electron microscope study. The general patterns of ultrastructure and the mode of cell division resemble Chroococcales. The cyanobacterium strain was axenic and cultured on both inorganic Z8 and BG13 media. Mammalian toxicity was confirmed by mice bioassay. The major sympton of poisoning was severe diarrhoea. Histopathological study shows a slight hepatotoxicosis associated with a pronounced change in the intestinal mucosa which shows swelling and destruction of villi epithelium and shedding of enterocytes into the lumen. Although slow, these kinds of poisoning are comparable to those induced by okadiac acid intraperitoneal mice injection (diarrhetic shellfish poisoning “DSP” toxins). By using the enzyme-linked immunosorbent assay (ELISA), the amount ofhepatotoxins “microcystins” was determined. The result shows that Synechocystis can produce a small amount of total microcystine [an average of 15 μg−1 dry weight corresponding to 20 ng(109cell)−1]. These findings lead us to consider Synechocystis as both a potent neurotoxin and hepatotoxin producer. Because of the confirmed cyanobacterium toxicity, an eventual ecological implication should be considered. However, a toxic chronic test experiment on Daphnia was simultaneously carried out. Juvenile D. magna (less than 24 hours old), were fed three concentrations (104, 106, 108 cells / ml) of Synechocystis. A group of organisms fed with Chlorella vulgaris (3. 105 cells/ml) and another group without food, were studied as control treatments. Only animals cultured with 104 cells/ml of cyanobacterium survived at 80% until the end of the test (21 days). Reproduction and normal growth occurred in control treatments fed with Chlorella vulgaris and the group fed with the lowest concentration of Synechocystis. One-way ANOVA statistical analyses show significant differences in Daphnia survival and growth, between treatments with and without Synechocystis and between treatments with and without food. In terms of this study, there is evidence that toxic picocyanobacteria blooms occurring in wastewater stabilization ponds of Marrakesh, could have harmful repercussions on zooplanktonic, bacteria and other algae communities. Consequently, this will constitute a possible hindrance for sewage self-purification process and system treatment performance. In addition, the reuse of such treated wastewater effluent for irrigation will constitute an additional, potent, health hazard for animals and human's.

Polarized Photoacoustic Spectra of the Cyanobacterium Synechococcus Oriented in Polymer Film

Photoacoustic spectra (PAS) were obtained for the cyanobacterium Synechococcus (Anacystis nidulans) cells embedded in isotropic and stretched polyvinyl alcohol films. The polarized radiation with the electric vector changing in 30° intervals with respect to given direction in a sample plane was used. Two cyanobacterium strains, one with very low biliprotein content, second with normal amount of biliproteins were investigated. The polarized absorption and fluorescence spectra were also measured. Conclusions were drawn about the thermal deactivation occurring in differently oriented pools of chromophores and about mutual orientation of their transition moments. Thermal deactivation in carotenoids (Cars) of both strains was different. The ratio of Car thermal deactivation to the thermal deactivation of chlorophyll (Chl) was higher in cyanobacteria with lower content of biliproteins than in the strain with normal amount of these complexes. Hence biliproteins can play the role in excitation energy transfer from Cars to Chls. For complex biological samples, polarized PAS can be a more sensitive method to investigate the directions of the absorption transition moments than the widely used polarized absorption spectra.

System Identification of Electronic Nose Data for Monitoring Cyanobacteria in Potable Water: Black-Box Modeling

Linear black-box modelling techniques are applied to data collected from an electronic nose experiment. FIR, ARX, MAX and ARMAX inverse models of the nose system are used to discriminate between two different strains of cyanobacteria. Models for both pre-processed and raw data are proposed, and successful classification rates of 100% and 99.3% are obtained, respectively.

Removal of nitrate from groundwater by cyanobacteria: quantitative assessment of factors influencing nitrate uptake.

The feasibility of biologically removing nitrate from groundwater was tested by using cyanobacterial cultures in batch mode under laboratory conditions. Results demonstrated that nitrate-contaminated groundwater, when supplemented with phosphate and some trace elements, can be used as growth medium supporting vigorous growth of several strains of cyanobacteria. As cyanobacteria grew, nitrate was removed from the water. Of three species tested, Synechococcus sp. strain PCC 7942 displayed the highest nitrate uptake rate, but all species showed rapid removal of nitrate from groundwater. The nitrate uptake rate increased proportionally with increasing light intensity up to 100 micromol of photons m(-2) s(-1), which parallels photosynthetic activity. The nitrate uptake rate was affected by inoculum size (i.e., cell density), fixed-nitrogen level in the cells in the inoculum, and aeration rate, with vigorously aerated, nitrate-sufficient cells in mid-logarithmic phase having the highest long-term nitrate uptake rate. Average nitrate uptake rates up to 0.05 mM NO(3-) h(-1) could be achieved at a culture optical density at 730 nm of 0.5 to 1. 0 over a 2-day culture period. This result compares favorably with those reported for nitrate removal by other cyanobacteria and algae, and therefore effective nitrate removal from groundwater using this organism could be anticipated on large-scale operations.

Effects of sulfonylurea herbicides on non-target aquatic micro-organisms: Growth inhibition of micro-algae and short-term inhibition of adenine and thymidine incorporation in periphyton communities

To study the differential sensitivity to sulfonylurea herbicides of aquatic micro-organisms we used the micro-plate technique as previously described by Blanck and Björnsäter [Blanck, H., Björnsäter, B., 1989. The algal microtest battery—a manual for routine tests of growth inhibition. Report to the Swedish National Chemicals Inspectorate, no 3/89]. This technique enables growth-inhibition tests to be carried out for a large number of species of micro-algae, and growth inhibition data for the two sulfonylureas chlorsulfuron and metsulfuron methyl were obtained for 20 marine species and 20 fresh-water species, respectively. The species-dependent variation in algal sensitivity was found to be very large, with strains of cyanobacteria and dinoflagellates being the most sensitive. The most sensitive species had EC 50 values for the inhibition of growth in the nanomolar range while EC 50 was higher than 1000 μM for the most tolerant species. Field experiments were also carried out to examine the sensitivity of periphyton communities in an agricultural area in southern Sweden. Periphyton communities were established on an artificial substratum in a creek affected by the runoff of agricultural pesticides and a creek with a watershed with no use of pesticides. Short-term toxicity of the sulfonylurea tribenuron methyl was assayed as inhibition of adenine and thymine incorporation weekly for 8 weeks at all sites. At the site with high exposure to pesticides and fertilisers the lowest observed effect concentration (LOEC) values for inhibition of adenine incorporation were always higher than environmentally expected concentrations indicating a high tolerance of periphyton to tribenuron methyl at this site. The toxicity of tribenuron methyl at the reference site was very variable over time. Adenine incorporation in periphyton from the reference site was in three out of seven experiments significantly inhibited at 3 nM tribenuron methyl, but in the remaining four experiments, LOEC values were much higher. The apparent high background variability in the short-term toxicity of tribenuron methyl makes an interpretation of the observed differences between the reference site and the site with higher exposure to pesticides very difficult. It is never the less interesting to compare the growth inhibition test data with some of the field toxicity tests at the reference site, showing that sulfonylurea herbicides can adversely affect both algal growth and periphyton activity at nanomolar concentrations.

Strategies of thermal adaptation by high‐latitude cyanobacteria

Although mat‐forming cyanobacteria dominate many freshwater ecosystems in the Arctic and Antarctic, their optimal temperature for growth (Topt) is usually much higher than the temperature range of their native habitat. The present study compared the temperature dependence of growth, pigment composition and absorbance, photosynthesis and photosynthate partitioning for two strains of cyanobacteria with contrasting Topt values; Phormidium subfuscum, isolated from McMurdo Ice Shelf, Antarctica, and Phormidium tenue, collected from the Kuparuk River in the tundra region of northern Alaska. Phormidium subfuscum grew between 5 and 20°C with a Topt of 15°C whereas P. tenue showed detectable growth from 10 to 40°C and a Topt of 30°C. Light utilization efficiency, photosynthetic capacity and the irradiance at the onset of light saturation increased with increasing temperature up to Topt in both strains. The cellular concentrations of chlorophyll a (Chl a) and carotenoid (CAR) and the in vivo absorbance maxima for Chl a, CAR, C‐phycocyanin and allophycocyanin changed little for P. subfuscum but all these variables increased across the temperature range up to Topt for P. tenue. Neither P. subfuscum nor P. tenue showed changes in relative carbon allocation with varying temperature, suggesting that gross biochemical alterations are not a characteristic of temperature acclimation in these cyanobacteria. We conclude that the eurythermal cyanobacterium P. tenue optimizes growth over a wide range of temperatures by adjusting its light‐capturing as well as carbon fixation characteristics, whereas stenothermal P. subfuscum relies on changes in carbon fixation without concomitant shifts in pigment content.

Response of two strains of Nostoc muscorum to metal stress and salinity

SummaryTwo strains of a cyanobacterium Nostoc muscorum, wild‐type N. muscorum (Cds) and an isolate having resistance to the heavy metal cadmium (Cdr), were selected for characterisation of their growth potential and physiological assays in certain defined stress environments.The chosen determinants were copper (heavy metal) and NaCl (salt stress). The observations on growth, heterocyst frequency, chlorophyll and nucleic acid contents, photosynthetic O2 evolution, 14C incorporation and acetylene reduction suggested that the strain Cdr was also resistant to copper. This strain, however, was found to be more sensitive to NaCl in comparison to its wild‐type counterpart. NaCl was found to enhance sugar accumulation in Cds and was more inhibitory to acetylene reduction rates than to the photosynthetic activities. The interaction between Cu and NaCl appeared to be antagonistic as the depression of growth and physiological activities by a mixture of the two was lesser than that caused by either of these. These observations form the first report on the response of a metal resistant strain of cyanobacterium to salinity.

Production, extraction and purification of phycobiliproteins from Nostoc sp.

One strain of nitrogen-fixing cyanobacterium ( Nostoc sp. PCC 9202) was grown indoors in 1 litre glass air-lift reactors as well as 17 litre polyethylene bags. Temperature dependence of growth-kinetic parameters, as well as of biomass and phycobiliprotein productivities, were determined. Harvesting and phycobiliprotein extraction methodologies are presented. The purification of crude extracts was performed by means of ultrafiltration or (NH 4) 2SO 4 precipitation followed by gel filtration and ion-exchange chromatography. Final phycocyanin and phycoerythrin preparations were characterized by purity ratios above 4 and 5, respectively. Phycobiliprotein production cost determination for each purity grade and cumulative phycobiliprotein weight losses along purification steps are shown. Production cost for high purity phycoerythrin (30 US$ per g) seems to be far below the market price.

First report of microcystins in Taiwan

This is the first report on microcystins from Microcystis aeruginosa Ku ̈tzing in Taiwan. A total of nine strains of cyanobacteria have been isolated from eutrophic aquaculture ponds and water reservoirs. By mouse toxicity assay, six of the nine strains had ld 100 in the range of 25–100 mg per kg mouse for dried bacterial mass. Microcystin-LR and -RR were found in all toxic strains and their contents ranged from 0.11–10.06 μg and 0.08–2.21 μg per mg of dried bacteria, respectively. Microcystin-RA, a minor component found only in M. TN-2 and M. CY-1 strains, was identified as a new microcystin. All three toxins were isolated by a serial separation on an LH-20 column, Si-flash column chromatography and reverse phase HPLC. Toxins were further identified by comparing their FABMS, 1H and 1H– 1H COSY NMR spectra with the authentic microcystin-LR. Several other microcystin-like compounds were also found in the cultured strains and their structures are being determined.

Determination of some physicochemical parameters of microcystins (cyanobacterial toxins) and trace level analysis in environmental samples using liquid chromatography

Some physicochemical parameters of three microcystin standards, known to be potent hepatotoxins produced by cyanobacteria, were determined using well defined chromatographic measurements. The logarithm of their retention factor on octadecylsilica (ODS) with water as the eluent, which is an estimation of the hydrophobicity of a molecule, was assessed at pH 7 at 3.9, 4.2 and 4.4 for microcystins-YR, -LR and -RR, respectively. Though being rather hydrophobic, microcystins also possess polar functions, namely carboxylic acids, amino and amido groups. The ionization of carboxylic groups occurs at pH values of 3.3–3.4. In environmental waters, microcystins are neutral or anionic. They are readily soluble in water, the solubility of microcystin-LR being higher than 1 g/l. Owing to their hydrophobicity and their polar functions, microcystins remain in the aqueous phase rather than being adsorbed on sediments or on suspended particulate matter. In a river water spiked with microcystins at 5 μg/l, only 10% was adsorbed on particles and 7% on the sandy sediment after three days. A method using solid-phase extraction on ODS followed by high-performance – or micro – liquid chromatography was optimized to detect microcystins at trace level in water. A clean-up was introduced to eliminate part of the interfering compounds coextracted during the sample percolation. Good recoveries (75–80%) were obtained. The method was linear, reproducible (with relative standard deviations ranging from 5 to 8%) and enabled the determination of microcystins at levels as low as 30 ng/l in drinking water and 100–200 ng/l in surface waters. The production of toxins by two strains of cyanobacteria was evaluated. Variations in the microcystin-LR content in the cells and in the medium of Microcystis aeruginosa PCC7806 were recorded over a five-week period. Toxin production was not correlated to the biomass but depended on the growth stage and was maximal at the end of the exponential growth phase. The release of toxin in water occurred essentially in old cultures where microcystin-LR was determined at concentrations of 170 and 280 μg/l in the media of M. aeruginosa PCC7806 and M. aeruginosa PCC7813, respectively. Other microcystins are likely to be synthesized by these strains. But owing to the lack of standards, mass spectrometric detection is required for further identification. This study points out the need of having other standards for water quality monitoring.

Expression of the mouse metallothionein-I gene conferring cadmium resistance in a transgenic cyanobacterium.

This paper reports the construction of a transgenic strain of cyanobacterium aimed at removing heavy metal pollution in waters. The mouse metallothionein-I (mMT-I) gene was inserted in the vector pRL-439 downstream of the strong psbA promoter. The resulting plasmid pRL-MT was ligated at the EcoRI site of the shuttle vector pKT-210 to generate the shuttle expression vector pKT-MT. This recombinant plasmid was introduced into Anabaena sp. PCC 7120 by triparental conjugative transfer. After selection on streptomycin, a stable transgenic Anabaena strain was obtained. The presence of the mMT-I gene was confirmed by DNA/DNA hybridization and its expression was demonstrated by immunodetection with specific antibodies. A metal tolerance experiment showed that this transgenic Anabaena strain had acquired higher metal resistance.

Expression of the mouse metallothionein-I gene conferring cadmium resistance in a transgenic cyanobacterium

This paper reports the construction of a transgenic strain of cyanobacterium aimed at removing heavy metal pollution in waters. The mouse metallothionein-I (mMT-I) gene was inserted in the vector pRL-439 downstream of the strong psbA promoter. The resulting plasmid pRL-MT was ligated at the EcoRI site of the shuttle vector pKT-210 to generate the shuttle expression vector pKT-MT. This recombinant plasmid was introduced into Anabaena sp. PCC 7120 by triparental conjugative transfer. After selection on streptomycin, a stable transgenic Anabaena strain was obtained. The presence of the mMT-I gene was confirmed by DNA/DNA hybridization and its expression was demonstrated by immunodetection with specific antibodies. A metal tolerance experiment showed that this transgenic Anabaena strain had acquired higher metal resistance.

Pp1-cyano2, a protein serine/threonine phosphatase 1 gene from the cyanobacterium Microcystis aeruginosa UTEX 2063.

Polymerase chain reaction (PCR) products similar to protein serine/threonine family I phosphatase genes were identified in five strains of cyanobacteria from three species. The gene for one of these protein phosphatase PCR products, pp1-cyano2 from Microcystis aeruginosa UTEX 2063, was cloned and sequenced. The deduced protein sequence PP1-cyano2 contains 264 amino acid residues ( approximately 30.3 kDa). In its N-terminal region, PP1-cyano2 had a GDXXHG(X)nGDXXDRG(X)nGNHE (nP23) sequence that is well-conserved in all protein serine/threonine family I phosphatases. Of 19 amino acid residues important for either metal binding, structure of the active site, or catalysis in eukaryotic PP1, 18 were present in PP1-cyano2. Reverse-transcription-PCR results showed that pp1-cyano2 was expressed under laboratory culture conditions.

New acylated sulfoglycolipids and digalactolipids and related known glycolipids from cyanobacteria with a potential to inhibit the reverse transcriptase of HIV-1.

Five novel diacylated sulfoglycolipids (1-5) were isolated from the cyanobacterium Scytonema sp. (TAU strain SL-30-1-4) and four novel acylated diglycolipids (6-9) were isolated from the cyanobacterium Oscillatoria raoi (TAU strain IL-76-1-2). These two groups of glycolipids and related known glycolipids isolated from these two and three other strains of cyanobacteria, Phormidium tenue (TAU strain IL-144-1), O. trichoides (TAU strain IL-104-3-2), and O. limnetica (TAU strain NG-4-1-2), were found to inhibit HIV-1 RT enzymatic activity to different extents. The structure elucidation of the various compounds is based on the selective hydrolysis of the glycerol ester moieties, GCMS analysis of the methyl ester derivatives of the liberated fatty acids, homo- and heteronuclear-2D-NMR techniques, and MS. The use of negative-ion FABMS for analyzing the combination and distribution of the fatty acids in glycolipids is demonstrated.

Cultural and molecular characterization of photobionts of Peltigera Membranacea

A molecular study was undertaken to clarify the identity of the photobiont in colourmorphs of the lichen, Peltigera membranacea. Two strains of cyanobacteria, identified as Nosroc sp. by morphology, were cultivated from each of two lichen specimens. Prokaryotic (16S) ribosomal RNA gene fragments were amplified by the polymerase chain reaction (PCR) from DNA extracted from the isolated strains and the lichens, and sequenced directly. Sequences were 98 1% identical between lichen specimens, TDI#AR94 and TDI#AR95, and highly similar to sequences published, or generated in this study from a type culture, for Nostoc. The 16S ribosomal RNA gene sequences (‘ 16S rDNA’) of all four lichen-derived cyanobacteria appeared the same, even though the lichen specimens from which they originated had different sequences. The 16S rDXA from strains 9A and 9B were different from that of specimen TDI#AR94, the thallus from which they were isolated, and instead were the same as that of strains 10A and 10B, and their source, specimen TDI#AR95. When primers selective for the strain 9A sequence were used, however, a small amount of PCR product corresponding to the 16S rDNA of strain 9A was obtained from lichen TDI#AR94. The results confirm that the photobionts of P. membranacea belong to Nostoc, and suggest that genetic differences in the photobiont may be a factor in the occurrence of colourmorphs among cyanolichens.