Cyanobacterium Leptolyngbya Research Articles

A Preliminary Report on the Aerophytic Cyanoprokaryota and Algae from the Great Smoky Mountains National Park, USA

An annotated list is provided for 38 species of aerophytic algae (31 cyanobacteria, six Chlorophyta, and one Bacillariophyta) that were recovered from eight soil study sites (in coniferous and hardwood forests) collected during surveys carried out in the Great Smoky Mountains National Park (GSMNP) in August 2010. The cyanobacterium Leptolyngbya foveolarum was the single most frequently encountered species recovered from all eight study areas. Most species (68% of the total) were found at only one or two sampling sites. There were 32 new species records in the GSMNP.

In vivo transposon tagging in the nonheterocystous nitrogen-fixing cyanobacterium Leptolyngbya boryana.

Nitrogenase is an oxygen-vulnerable metalloenzyme that catalyzes nitrogen fixation. It largely remains unknown how nitrogenase coexists with oxygenic photosynthesis in nonheterocystous cyanobacteria, since there have been no appropriate model cyanobacteria so far. Here, we demonstrate in vivo transposon tagging in the nonheterocystous cyanobacterium Leptolyngbya boryana as a forward genetics approach. By conjugative transfer, a mini-Tn5-derived vector, pKUT-Tn5-Sm/Sp, was transferred from Escherichia coli to L. boryana cells. Of 1839 streptomycin-resistant colonies, we isolated three mutants showing aberrant diazotrophic growth. Genome resequencing identified the insertion sites of the transposon in the mutants. This in vivo transposon tagging mutagenesis of L. boryana provides a promising system to investigate molecular mechanisms to resolve the Oxygen Paradox between nitrogen fixation and oxygenic photosynthesis in cyanobacteria.

A Leptolyngbya-based microbial consortium for agro-industrial wastewaters treatment and biodiesel production

A mixed cyanobacterial-mixotrophic algal population, dominated by the filamentous cyanobacterium Leptolyngbya sp. and the microalga Ochromonas (which contributed to the total photosynthetic population with rates of less than 5%), was studied under non-aseptic conditions for its efficiency to remove organic and inorganic compounds from different types of wastes/wastewaters while simultaneously producing lipids. Second cheese whey, poplar sawdust, and grass hydrolysates were used in lab-scale experiments, in photobioreactors that operated under aerobic conditions with different initial nutrient (C, N and P) concentrations. Nutrient removal rates, biomass productivity, and the maximum oil production rates were determined. The highest lipid production was achieved using the biologically treated dairy effluent (up to 14.8% oil in dry biomass corresponding to 124mgL-1) which also led to high nutrient removal rates (up to 94%). Lipids synthesized by the microbial consortium contained high percentages of saturated and mono-unsaturated fatty acids (up to 75% in total lipids) for all the substrates tested, which implies that the produced biomass may be harnessed as a source of biodiesel.

Morphophysiological and transcriptome analysis reveals a multiline defense system enabling cyanobacterium Leptolyngbya strain JSC-1 to withstand iron induced oxidative stress

Iron intoxications induce severe oxidative stress by producing reactive oxygen species (ROS) in cyanobacteria, leading to membrane lipid peroxidation, altered morphology, impaired photosynthesis and other oxidative stress injuries. Given these stresses, mitigation of ROS is a prerequisite for all aerobic organisms. Study of siderophilic cyanobacterium Leptolyngbya strain JSC-1 inhabiting iron-rich hot springs may provide insight into the mechanism of iron homeostasis and alleviation of oxidative stress. In this study, we investigated the morphophysiological and molecular mechanisms enabling this cyanobacterium to cope with iron-induced oxidative stress. Strain JSC-1 biomineralized extracellular iron via an exopolymeric sheath (acting as a first line of defense) and intracellular iron via polyphosphate inclusions (second line of defense), thus minimizing the burden of free ferric ions. Physiological parameters, SOD, CAT and POD activities, bacterioferritin and total protein contents fluctuated in response to iron elevation, displaying a third line of defense to mitigate ROS. Differential gene expression analysis of JSC-1 indicated up-regulation of 94 and 125 genes and down-regulation of 89 and 183 genes at low (4 μM) and high (400 μM) iron concentration, respectively. The differentially expressed genes (DEGs) were enriched in 100 KEGG pathways and were found to be involved in lipopolysaccharide and fatty acid biosynthesis, starch, sucrose, chlorophyll and other metabolic pathways. Together with metabolic reprogramming (fourth line of defense), JSC-1 established a unique multiline defense system that allows JSC-1 to withstand severe oxidative stress. These findings also provide insight into potential survival strategies of ancient microorganisms inhabiting similar environment present in early earth history.

Biofabrication of gold nanoparticles by Lyptolyngbya JSC-1 extract as super reducing and stabilizing agents: Synthesis, characterization and antibacterial activity

This study describes the first ever utilization of cell free aqueous extract of cyanobacterium Leptolyngbya JSC-1 as a source of strong reducing and stabilizing agents for the optimal biofabrication of gold nanoparticles (AuNPs) through an eco-friendly synthetic route. Well dispersed crystalline AuNPs of spherical morphology with a particle size of 100–200 nm were prepared. FTIR spectral analysis was then performed to characterize the possible functionalities of JSC-1 extract, mainly involved in stabilizing and formation of AuNPs. Based on the redox potential of JSC-1 extract, it was further confirmed that the extract provide a strong reducing environment in the reaction medium and causes reduction of gold ions. The resultant AuNPs were then explored to find out their photo-catalytic activity for methylene blue and antibacterial activities against E. coli (18 ± 2 mm) and S. aureus (14 ± 2 mm). It has been mechanistically identified that AuNPs caused bacterial membrane damage and cell disruption by inducing the production of intracellular reactive oxygen species (ROS). Together, these finding reveals that biochemically capped AuNPs are the promising antibacterial agents that induce oxidative stress in the two bacterial species evaluated and cause their membrane disruption leading to cell leakage and death.

Biocrust ecology: unifying micro- and macro-scales to confront global change.

Biocrust ecology: unifying micro- and macro-scales to confront global change.

Leptolyngbyolides, Cytotoxic Macrolides from the Marine Cyanobacterium Leptolyngbya sp.: Isolation, Biological Activity, and Catalytic Asymmetric Total Synthesis.

Four new macrolactones, leptolyngbyolides A-D, were isolated from the cyanobacterium Leptolyngbya sp. collected in Okinawa, Japan. The planar structures of leptolyngbyolides were determined by extensive NMR studies, although complete assignment of the absolute configuration awaited the catalytic asymmetric total synthesis of leptolyngbyolide C. The synthesis took advantage of the catalytic asymmetric thioamide-aldol reaction using copper(I) complexed with a chiral bidentate phosphine ligand to regulate two key stereochemistries of the molecule at the outset. The present total synthesis demonstrates the utility of this reaction for the construction of complex chemical entities. In addition to the total synthesis, this work reports that leptolyngbyolides depolymerize filamentous actin (F-actin) both in vitro and in cells. Detailed biological studies suggest the probable order of F-actin depolymerization and apoptosis caused by leptolyngbyolides.

The Effect of Two Amino acid Residue Substitutions via RNA Editing on Dark-operative Protochlorophyllide Oxidoreductase in the Black Pine Chloroplasts

Dark-operative protochlorophyllide oxidoreductase (DPOR) is a key enzyme to produce chlorophyll in the dark. Among photosynthetic eukaryotes, all three subunits chlL, chlN, and chlB are encoded by plastid genomes. In some gymnosperms, two codons of chlB mRNA are changed by RNA editing to codons encoding evolutionarily conserved amino acid residues. However, the effect of these substitutions on DPOR activity remains unknown. We first prepared cyanobacterial ChlB variants with amino acid substitution(s) to mimic ChlB translated from pre-edited mRNA. Their activities were evaluated by measuring chlorophyll content of dark-grown transformants of a chlB-lacking mutant of the cyanobacterium Leptolyngbya boryana that was complemented with pre-edited mimic chlB variants. The chlorophyll content of the transformant cells expressing the ChlB variant from the fully pre-edited mRNA was only one-fourth of the control cells. Co-purification experiments of ChlB with Strep-ChlN suggested that a stable complex with ChlN is greatly impaired in the substituted ChlB variant. We then confirmed that RNA editing efficiency was markedly greater in the dark than in the light in cotyledons of the black pine Pinus thunbergii. These results indicate that RNA editing on chlB mRNA is important to maintain appropriate DPOR activity in black pine chloroplasts.

CO2 Removal from Biogas by Cyanobacterium Leptolyngbya sp. CChF1 Isolated from the Lake Chapala, Mexico: Optimization of the Temperature and Light Intensity.

In the present study, the capacity of the cyanobacterium Leptolyngbya sp. CChF1 to remove CO2 from real and synthetic biogas was evaluated. The identification of the cyanobacterium, isolated from the lake Chapala, was carried out by means of morphological and molecular analyses, while its potential for CO2 removal from biogas streams was evaluated by kinetic experiments and optimized by a central composite design coupled to a response surface methodology. Results demonstrated that Leptolyngbya sp. CChF1 is able to remove CO2 and grow indistinctly in real or synthetic biogas streams, showing tolerance to high concentrations of CO2 and CH4, 25 and 75%, respectively. The characterization of the biomass composition at the end of the kinetic assays revealed that the main accumulated by-products under both biogas streams were lipids, followed by proteins and carbohydrates. Regarding the optimization experiments, light intensity and temperature were the studied variables, while synthetic biogas was the carbon source. Results showed that light intensity was significant for CO2 capture efficiency (p=0.0290), while temperature was significant for biomass production (p=0.0024). The predicted CO2 capture efficiency under optimal conditions (27.1°C and 920lx) was 93.48%. Overall, the results of the present study suggest that Leptolyngbya sp. CChF1 is a suitable candidate for biogas upgrading.

Biotreatment of raisin and winery wastewaters and simultaneous biodiesel production using a Leptolyngbya-based microbial consortium

Combining wastewater treatment with biodiesel production can greatly reduce the cost of environmentally friendly technologies. In this work wastewaters of high environmental importance originating from raisin and wine industries were employed as substrates for a mixed cyanobacterial/algal cultivation system dominated by the filamentous cyanobacterium Leptolyngbya sp. The mixotrophic microalga Ochromonas contributed to less than 5% of the total photosynthetic population and grazed on broken Leptolyngbya trichomes. Satisfactory lipid yields and removal of nutrients and chemical oxygen demand (COD) were observed when raisin or winery wastewaters were used separately as substrates. However, exceptional removal rates were recorded in blends of the above-mentioned wastewaters (i.e., 92.8%, 78.1% and 99% for COD, total nitrogen and phosphate, respectively), while the biomass produced contained around 13% lipids (w/w) on a dry weight basis. The ratio of saturated/mono-unsaturated fatty acids in this lipid reached 85%, making this system suitable for biodiesel production. The proposed system can effectively approach a technology that integrates wastewater treatment with simultaneous biodiesel production.

Therapeutic properties in Tunisian hot springs: first evidence of phenolic compounds in the cyanobacterium Leptolyngbya sp. biomass, capsular polysaccharides and releasing polysaccharides.

BackgroundIn Tunisia, the use of hot spring waters for both health and recreation is a tradition dating back to Roman times. In fact, thermal baths, usually called “Hammam” are recommended as a therapeutic and prophylactic measure against many types of illness and toxicity. While the chemical concentration of thermal water is admittedly associated with its therapeutic effects, the inclusion in spa waters of efficient bioproduct additives produced by photosynthetic microorganisms and that act against oxidative stress may comprise a significant supplementary value for thermal centers. The aim of this study was to investigate the antioxidant potential of the Tunisian thermophilic cyanobacterium Leptolyngbya sp. and to determine its phytochemical constituents and phenolic profile.MethodsBME (Biomass Methanolic Extract), CME (Capsular polysaccharides Methanolic Extract) and RME (Releasing polysaccharides Methanolic Extract) of Leptolyngbya sp. were examined for their antioxidant activities by means of DPPH, hydroxyl radical scavenging and ferrous ion chelating assays. Their total phenols, flavonoids, carotenoids, Mycosporine-like amino acids (MAAs) and vitamin C contents, as well as their phenolic profiles were also determined.ResultsBME has the highest content of phenols (139 ± 1.2 mg/g), flavonoids (34.9 ± 0.32 mg CEQ/g), carotenoids (2.03 ± 0.56 mg/g) and vitamin C (15.7 ± 1.55 mg/g), while the highest MAAs content (0.42 ± 0.03 mg/g) was observed in CME. BME presented both the highest DPPH and hydroxyl radical scavenging ability with an IC50 of 0.07 and 0.38 mg/ml, respectively. The highest ferrous chelating capacity was detected in CME with an IC50 = 0.59 mg/ml. Phenolic profiles revealed the presence of 25 phenolic compounds with the existence of hydroxytyrosol, oleuropein, resveratrol and pinoresinol.ConclusionThe study demonstrated that the cyanobacterium Leptolyngbya sp. possesses abundant natural antioxidant products which may have prophylactic and therapeutic effects on many types of illness and toxicity. The present findings not only explain and reinforce the rationale behind traditional therapeutic practices in Tunisia in the exploitation of the country’s hot springs, but support the addition of Leptolyngbya to thermal waters as a means to enhance the value and reputation of the curative nature of Tunisian thermal waters.

Total synthesis and stereochemical determination of yoshinone A

In 2014, the γ-pyrone-containing polyketide, yoshinone A, was isolated from the marine cyanobacterium Leptolyngbya sp. and its structure was determined. Yoshinone A inhibited differentiation of 3T3-L1 cells into adipocytes, with an EC50 value of 420 nM without any cytotoxicity, and therefore is expected to be a lead compound for obesity drugs. To establish its absolute configuration, and to provide sufficient amounts for further research, the total synthesis of yoshinone A was achieved through synthesis of its two possible diastereomers.

Identification of a cis-acting element in nitrogen fixation genes recognized by CnfR in the nonheterocystous nitrogen-fixing cyanobacterium Leptolyngbya boryana.

The filamentous cyanobacterium Leptolyngbya boryana has the ability to fix nitrogen without any heterocysts under microoxic conditions. Previously, we identified the cnfR gene for a master transcriptional activator for nitrogen fixation (nif) genes in a 50-kb gene cluster containing nif and nif-related genes in L. boryana. We showed that CnfR activates the transcription of nif genes in response to low oxygen conditions, which allows the oxygen-vulnerable enzyme nitrogenase to function. However, the regulatory mechanism that underlies regulation by CnfR remains unknown. In this study, we identified a conserved cis-acting element that is recognized by CnfR. We established a reporter system in the non-diazotrophic cyanobacterium Synechocystis sp. PCC 6803 using luciferase genes (luxAB). Reporter analysis was performed with a series of truncated and modified upstream regulatory regions of nifB and nifP. The cis-element can be divided into nine motifs I-IX, and it is located 76 bp upstream of the transcriptional start sites of nifB and nifP. Six motifs of them are essential for transcriptional activation by CnfR. This cis-acting element is conserved in the upstream regions of nif genes in all diazotrophic cyanobacteria, including Anabaena and Cyanothece, thereby suggesting that the transcriptional regulation by CnfR is widespread in nitrogen-fixing cyanobacteria.

Inhibition of Vibrio spp. by 2‐Hydroxyethyl‐11‐hydroxyhexadec‐9‐enoate of Marine Cyanobacterium Leptolyngbya sp. LT 19

Inhibition of <i>Vibrio</i> spp. by 2‐Hydroxyethyl‐11‐hydroxyhexadec‐9‐enoate of Marine Cyanobacterium <i>Leptolyngbya</i> sp. <scp>LT</scp> 19

The siderophilic cyanobacterium Leptolyngbya sp. strain JSC-1 acclimates to iron starvation by expressing multiple isiA-family genes.

In the evolution of different cyanobacteria performing oxygenic photosynthesis, the core complexes of the two photosystems were highly conserved. However, cyanobacteria exhibit significant diversification in their light-harvesting complexes and have flexible regulatory mechanisms to acclimate to changes in their growth environments. In the siderophilic, filamentous cyanobacterium, Leptolyngbya sp. strain JSC-1, five different isiA-family genes occur in two gene clusters. During acclimation to Fe limitation, relative transcript levels for more than 600 genes increased more than twofold. Relative transcript levels were ~250 to 300 times higher for the isiA1 gene cluster (isiA1-isiB-isiC), and ~440- to 540-fold for the isiA2-isiA3-isiA4-cpcG2-isiA5 gene cluster after 48h of iron starvation. Chl-protein complexes were isolated and further purified from cells grown under Fe-replete and Fe-depleted conditions. A single class of particles, trimeric PSI, was identified by image analysis of electron micrographs of negatively stained PSI complexes from Fe-replete cells. However, three major classes of particles were observed for the Chl-protein supercomplexes from cells grown under iron starvation conditions. Based on LC-MS-MS analyses, the five IsiA-family proteins were found in the largest supercomplexes together with core components of the two photosystems; however, IsiA5 was not present in complexes in which only the core subunits of PSI were detected. IsiA5 belongs to the same clade as PcbC proteins in a phylogenetic classification, and it is proposed that IsiA5 is most likely involved in supercomplexes containing PSII dimers. IsiA4, which is a fusion of an IsiA domain and a C-terminal PsaL domain, was found together with IsiA1, IsiA2, and IsiA3 in complexes with monomeric PSI. The data indicate that horizontal gene transfer, gene duplication, and divergence have played important roles in the adaptive evolution of this cyanobacterium to iron starvation conditions.

A Collaborative Evaluation of LC-MS/MS Based Methods for BMAA Analysis: Soluble Bound BMAA Found to Be an Important Fraction.

Exposure to β-N-methylamino-l-alanine (BMAA) might be linked to the incidence of amyotrophic lateral sclerosis, Alzheimer’s disease and Parkinson’s disease. Analytical chemistry plays a crucial role in determining human BMAA exposure and the associated health risk, but the performance of various analytical methods currently employed is rarely compared. A CYANOCOST initiated workshop was organized aimed at training scientists in BMAA analysis, creating mutual understanding and paving the way towards interlaboratory comparison exercises. During this workshop, we tested different methods (extraction followed by derivatization and liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS) analysis, or directly followed by LC-MS/MS analysis) for trueness and intermediate precision. We adapted three workup methods for the underivatized analysis of animal, brain and cyanobacterial samples. Based on recovery of the internal standard D3BMAA, the underivatized methods were accurate (mean recovery 80%) and precise (mean relative standard deviation 10%), except for the cyanobacterium Leptolyngbya. However, total BMAA concentrations in the positive controls (cycad seeds) showed higher variation (relative standard deviation 21%–32%), implying that D3BMAA was not a good indicator for the release of BMAA from bound forms. Significant losses occurred during workup for the derivatized method, resulting in low recovery (<10%). Most BMAA was found in a trichloroacetic acid soluble, bound form and we recommend including this fraction during analysis.

Kalkipyrone B, a marine cyanobacterial γ-pyrone possessing cytotoxic and anti-fungal activities

Bioassay-guided fractionation of two marine cyanobacterial extracts using the H-460 human lung cancer cell line and the OVC-5 human ovarian cancer cell line led to the isolation of three related α-methoxy-β, β′-dimethyl-γ-pyrones each containing a modified alkyl chain, one of which was identified as the previously reported kalkipyrone and designated kalkipyrone A. The second compound was an analog designated kalkipyrone B. The third was identified as the recently reported yoshinone A, also isolated from a marine cyanobacterium. Kalkipyrone A and B were obtained from a field-collection of the cyanobacterium Leptolyngbya sp. from Fagasa Bay, American Samoa, while yoshinone A was isolated from a field-collection of cyanobacteria (cf. Schizothrix sp.) from Panama. One-dimensional and two-dimensional NMR experiments were used to determine the overall structures and relative configurations of the kalkipyrones, and the absolute configuration of kalkipyrone B was determined by 1H NMR analysis of diastereomeric Mosher’s esters. Kalkipyrone A showed good cytotoxicity to H-460 human lung cancer cells (EC50=0.9μM), while kalkipyrone B and yoshinone A were less active (EC50=9.0μM and >10μM, respectively). Both kalkipyrone A and B showed moderate toxicity to Saccharomyces cerevisiae ABC16-Monster strain (IC50=14.6 and 13.4μM, respectively), whereas yoshinone A was of low toxicity to this yeast strain (IC50=63.8μM).

Wastewater treatment and microbial communities in an integrated photo-bioelectrochemical system affected by different wastewater algal inocula

Integrated photo-bioelectrochemical (IPB) systems are a newly emerging technology for sustainable wastewater treatment through synergistic cooperation between microbial fuel cells (MFCs) and algal bioreactors. This study aimed to advance our understanding of the IPB performance in contaminant removal and bioenergy production and how IPB function is affected by different algal–bacterial inocula. Fed with a synthetic solution, the IPB system could achieve more than 90% removal of solution organic compounds and nearly 100% of ammonium nitrogen. Production of algal biomass was significantly different among three inocula, varying from 5.9 to 53.3mgL−1. The highest energy production was 0.089kWhm−3, including direct electrical energy of 0.055kWhm−3 and indirect electrical energy of 0.034kWhm−3 from biomass conversion, and a positive energy balance could be achieved. The natural algal inocula resulted in the cyanobacteria Leptolyngbya and green alga Acutodesmus as dominant photoautotrophs in cathode suspension and biofilms, providing oxygen for MFC function. Differences in IPB efficiency could be related to microbial composition; one inoculum resulted in absence of Xanthomonadaceae bacteria, while another had more γ-proteobacteria. Specific taxa identified could be important for optimizing electricity generation and algal biomass for biofuel production.

A first report of biodeterioration caused by cyanobacterial biofilms of exposed fossil bones: A case study of the middle Pleistocene site of La Polledrara di Cecanibbio (Rome, Italy)

La Polledrara di Cecanibbio is a Pleistocene fossiliferous deposit near Rome (Italy) where large quantities of bones, belonging mainly to elephants, have been discovered and ‘musealized’ under an enclosing structure. The prevailing environmental conditions inside the museum and the exposition in situ of the fossilized remains have resulted in the development of phototrophic biofilms on the bones and the nearby sediments. Samples of bones and sediment were investigated with different microscopy techniques (light, confocal laser scanning, scanning electron and petrographic microscopes) that allowed the identification of the cyanobacterium Leptolyngbya sp. as the almost entirely dominant organism of the biofilms. The present study identifies for the first time endolithic activity of this cyanobacterium on exposed fossilized bones. There was some indication that this species was euendolithic but this remains to be elucidated. The development of these phototrophic biofilms greatly reduces the aesthetic value of the site and evidence suggests that they may cause extensive structural damage to the bones, threatening the scientific and cultural assets of one of the richest fossiliferous deposits in Italy. This study on the biodeterioration of fossil remains gives useful insights for the conservation of this spectacular site.

Thermogenic travertine deposits in Thermopylae hot springs (Greece) in association with cyanobacterial microflora

The present paper deals with the investigation of abiotic and biotic influence on thermogenic travertine formation in Thermopylae hot springs, one of the largest active thermogenic travertine systems in Greece. Geological, mineralogical and microbiological data from three different types of travertines (cascades, terraces and fluvial crusts) revealed different cyanobacterial communities. Microscopic analysis of fresh and cultured material has shown that epilithic and endolithic cyanobacteria are almost the exclusive components of travertines’ photosynthetic microflora. Thirty-one (31) taxa of cyanobacteria are presented here, among them the frequently found, in such environments, Phormidium incrustatum and Aphanocapsa thermalis, as well as the taxonomically interesting diazotrophic morphotype identified as Chlorogloeopsis sp. Sampling sites I and II have similar formation conditions characterized by laminated travertines with low porosity and shrub lithotypes, with the cyanobacterium Leptolyngbya ercegovicii occupying an endolithic zone, while the upper part is occupied by colonial chroococcalean species. On the contrary, sampling site III is characterized by laminated travertine with fenestrial type porosity and absence of shrub lithotypes resulting in a completely different community of cyanobacteria.