Investigations into the processes of sorption and desorption of polypeptide antibiotics on Taunit multiwalled carbon nanotubes

A modern nanomaterial made of Taunit multiwalled carbon nanotubes (MWCNTs) was used for the first time to adsorb and desorb gramicidin S and teicoplanin A2 polypeptide antibiotics. Taunit was found to absorb efficiently antibiotics under particular conditions when MWCNTs were taken in excess at 20°C for 18 h. The desorption efficiency depended on the eluent content and the structure of antibiotics. The teicoplanin A2 antibiotic was eluted with a higher yield because of its higher polarity compared to gramicidin S (86% against 65% of the total Taunit-bound antibiotic).     

A sensitive photosystem II-based biosensor for detection of a class of herbicides

We have developed a biosensor for the detection of residual triazine-, urea- and phenolic-type herbicides, using isolated photosystem II (PSII) particles from the thermophilic cyanobacterium, Synechococcus elongatus, as biosensing elements. The herbicide detection was based on the fact that, in the presence of artificial electron acceptors, the light-induced electron transfer through isolated PSII particles is accompanied by the release of oxygen, which is inhibited by the herbicide in a concentration-dependent manner. The PSII particles were immobilized between dialysis membrane and the Teflon membrane of the Clark oxygen electrode mounted in a flow cell that was illuminated. Inclusion of the antibiotic chloramphenicol in the reaction mixtures prolonged, by 50%, the lifetime of the biosensor. The use of highly active PSII particles in combination with the flow system resulted in a reusable herbicide biosensor with good stability (50% of initial activity was still remaining after 35-h use at 25 degrees C) and high sensitivity (detection limit for diuron was 5 x 10(-10) M).     

Postprandial variations in the activity of polysaccharide-degrading enzymes of fluid- and particle-associated ruminal microbial populations

Effect of an algicidal product fromOscillatoria late-virens and of the herbicide 3-(3,4-dichlorophenyl)-1,1-dimethyl urea (DCMU) on growth, photosynthesis, and mouse toxicity inMicrocystis PCC 7820 was examined. Their lethal concentrations abolished photosystem (PS)-II reactions and eventually bleached and detoxified the cyanobacterium. Although loss of protein, chlorophyll, and toxicity were also induced by sublethal antibiotic doses, photosynthetic activities remained unchanged and developed antibiotic tolerance. These effects could be duplicated in natural conditions, implying utility of the natural algicide in control of toxic cyanobacteria.     

Production of the antibiotic-algicide cyanobacterin LU-2 by a filamentous cyanobacterium Nostoc sp.]

A strain of cyanobacterium of Nostoc has been isolated, and found to produce a new antibiotic cyanobacterin LU-2. The antibiotic is synthesized by the cyanobnacterium under intensive cultivation conditions in a liquid mineral medium. Cyanobacterin LU-2 is an exometabolite; its maximum accumulation in the medium is achieved at 34 degrees. Cyanobacterin LU-2 is active against many cyanobacteria tested, including those of Microcystis and Aphanizomenon which are principals to give rise to blooms in fresh water supplies. It is poorly active against green algae and inactive against fungi and bacteria. The antibiotic hinders cell division in Synechococcus sp. R-2 (PCC 7942). It causes compression of the cytoplasm and exfoliation of the cell contents from cell wall; the distance between tylacoids is increased and their destruction is observed. The antibiotic hinders markedly light-dependent oxygen evolution. Cyanobacterin LU-2 is substance of a phenolic nature containing amino-sugar.     

Surface modifications based on the cyanobacterial siderophore anachelin: from structure to functional biomaterials design

This review describes the design, synthesis and evaluation of novel catechol based anchors for surface modification. The anachelin chromophore, the catecholate fragment of the siderophore anachelin from the cyanobacterium Anabaena cylindrica, allows for the immobilization of polyethylene glycol (PEG) on titania and glass surfaces thus rendering them protein resistant and antifouling. It is proposed that catecholate siderophores constitute a class of natural products useful for surface modification similar to dihydroxyphenylalanine and dopamine derived compounds found in mussel adhesive proteins. Second-generation dopamine derivatives featuring a quaternary ammonium group were found to be equally efficient in generating antifouling surfaces. The anachelin chromophore, merged via a PEG linker to the glycopeptide antibiotic vancomycin, allowed for the generation of antimicrobial surfaces through an operationally simple dip-and-rinse procedure. This approach offers an option for the prevention of nosocomial infections through antimicrobial implants, catheters and stents. Consequences for the mild generation of functional biomaterials are discussed and novel strategies for the immobilization of complex natural products, proteins and DNA on surfaces are presented.     

Effects of Photosystem II Herbicides on the Photosynthetic Membranes of the Cyanobacterium Aphanocapsa 6308

The effects of the photosystem II herbicides diuron (3-(3,4-dichlorophenyl)-1,1-dimethylurea) and atrazine (2-chloro-4-ethylamino-6-isopropylamino-s-triazine) on the photosynthetic membranes of a cyanobacterium, Aphanocapsa 6308, were compared to the effects on a higher plant, Spinacia oleracea. The inhibition of photosystem II electron transport by these herbicides was investigated by measuring the photoreduction of the dye 2,6-dichlorophenol-indophenol spectrophotometrically using isolated membranes. The concentration of herbicide that caused 50% inhibition of electron transport (I₅₀ value) in Aphanocapsa membranes for diuron was 6.8 × 10⁻⁹ molar and the I₅₀ value for atrazine was 8.8 × 10⁻⁸ molar. ¹⁴C-labeled diuron and atrazine were used to investigate herbicide binding with calculated binding constants (K) being 8.2 × 10⁻⁸ molar for atrazine and 1.7 × 10⁻⁷ molar for diuron. Competitive binding studies carried out on Aphanocapsa membranes using radiolabeled [¹⁴C]atrazine and unlabeled diuron revealed that diuron competed with atrazine for the herbicide-binding site. Experiments involving the photoaffinity label [¹⁴C]azidoatrazine (2-azido-4-ethylamino-6-isopropylamino-2-triazine) and autoradiography of polyacrylamide gels indicated that the herbicide atrazine binds to a 32-kilodalton protein in Aphanocapsa 6308 cell extracts.     

Fine-Tuning the Antimicrobial Profile of Biocompatible Gold Nanoparticles by Sequential Surface Functionalization Using Polyoxometalates and Lysine

Antimicrobial action of nanomaterials is typically assigned to the nanomaterial composition, size and/or shape, whereas influence of complex corona stabilizing the nanoparticle surface is often neglected. We demonstrate sequential surface functionalization of tyrosine-reduced gold nanoparticles (AuNPs^Tyr) with polyoxometalates (POMs) and lysine to explore controlled chemical functionality-driven antimicrobial activity. Our investigations reveal that highly biocompatible gold nanoparticles can be tuned to be a strong antibacterial agent by fine-tuning their surface properties in a controllable manner. The observation from the antimicrobial studies on a gram negative bacterium Escherichia coli were further validated by investigating the anticancer properties of these step-wise surface-controlled materials against A549 human lung carcinoma cells, which showed a similar toxicity pattern. These studies highlight that the nanomaterial toxicity and biological applicability are strongly governed by their surface corona.     

Biotransformation and biomonitoring of phenylurea herbicide diuron

A Gram-positive, Micrococcus sp. strain PS-1 isolated from diuron storage site was studied for its capability of biotransformation of phenylurea herbicide diuron to a secondary metabolite, 1-(3,4-dichlorophenyl)urea (DCPU) for bioconjugation and antibody development applications. The metabolite formed associated with profound changes in bacterial cell morphology demonstrated increase in the degradation kinetics of diuron in presence of small quantity of a surfactant. The synthesized metabolite identified by chromatographic and mass spectrometry techniques was conjugated with carrier protein, and used as an immunogen for antibodies production. The generated antibody was highly specific, demonstrating excellent sensitivity against diuron. The antibody was used as receptor molecules in standard fluorescence immunoassay (FIA) format showing detection limit of 0.01 ng/mL in the optimum working concentration range of diuron with good signal precision (～2%). The study presented first time the degradation pathway of herbicide by specific microorganism to synthesize hapten for bioconjugation and immunoassay development.     

Bioproduction of antimicrobial compounds by using marine filamentous cyanobacterium cultivation

The synthesis of bioactive compounds with antimicrobial activity, excreted by marine cyanobacteria, strongly depends on their growth conditions. Due to the wide variety of biomolecules which could show properties as growth inhibitors and their low concentrations within the culture medium, the activity of their crude extracts also seems to be related to the extraction method used. Using the marine filamentous cyanobacterium Geitlerinema strain Flo1, we demonstrate a systematic approach for identifying optimal culture conditions to obtain culture media extracts with antimicrobial activity. The changes in the culture conditions, such as the addition of NaBr to the medium, cell immobilisation in vegetable sponge pieces, or temperature, effected the production of these bioactive compounds. The crude extract, containing middle polar molecules, obtained by extraction with Amberlite XAD-1180 had a higher antifouling activity upon a number of bacteria and fungi than the extract obtained by extraction with Amberlite XAD-16. The lowest inhibitory concentration obtained upon Rhodosporidium sphaerocarpum was still ten times higher than that of bis(tributyltin)oxide, but compared to zinc pyrithione, it was two times more active.     

Construction of a non-antibiotic expression system in a marine cyanobacterium Synechococcus sp. PCC 7002 and its application in production of oral vaccine against enterotoxin of Escherichia coli

Synechococcus sp. PCC 7002 is a marine cyanobacterium that is rich in thylakoid membranes and performs oxygenic photosynthesis. In this study, we inactivated its psbEF genes encoding cytochrome b-559 of photosystem II and the mutant strain EF211 could not grow photoautotrophically. A non-antibiotic plasmid (pAQEF559) was constructed in such a way that it has a psbEF operon and all antibiotic genes are deleted. The plasmid could complement EF211 and restore photoautotrophical growth. This non-antibiotic plasmid was used to express the gene encoding heat-labile enterotoxin B-subunit, LT-B. The overproduced LT-B in the cyanobacterium could form pentamers and was associated with thylakoid membranes. Oral immunization of mice with the cyanobacterium overproducing LT-B showed that it was very effective in stimulating production of both IgG in serum and IgA in intestine. Our results demonstrate that the non-antibiotic vector in combination with the constructed host cyanobacterium is useful in production of recombinant proteins in cyanobacteria and the cyanobacterial cells producing LT-B can be an effective oral vaccine against enterotoxins.     

Herbicide effects on the population growth of some green algae and cyanobacteria

Six herbicides were tested for their effects on the population growth of a range of green algae and cyanobacteria by an easily replicated low-volume liquid culture technique using Repli-dishes. Diuron, propanil and atrazine were most inhibitory, chlorpropham was intermediate and MCPA and glyphosate were least inhibitory. Chlorpropham was more inhibitory to green algae than to cyanobacteria. The effects of chlorpropham and 3-chloroaniline, a metabolite, on populations of the cyanobacterium Anacystis nidulans and the alga Chlamydomonas reinhardii were monitored in larger scale batch cultures. Both compounds reduced the growth rate although in some cases there was partial recovery. 3-Chloroaniline was less inhibitory than the parent herbicide chlorpropham.     

Outdoor evaluation of herbicide resistant strains of Anabaena variabilis as biofertilizer for rice plants

Application of diazotrophic cyanobacteria, Anabaena variabilis, as biofertilizer for rice cultivation has a beneficial effect on crop productivity and maintenance of soil fertility. However, periodic applications of herbicides used to obtain high crop productivity are not only detrimental to weeds but to biofertilizer strains of cyanobacteria also. Therefore, research was undertaken to isolate four herbicide resistant strains (Arozin-R, Alachlor-R, Butachlor-R and 2,4-D-R) and a multiple herbicide resistant strain (MHR) of natural isolates of A. variabilis exhibiting resistance against these common rice field herbicides. The outdoor survivability of mutant strains and the productivity of rice crop (IR-36) were evaluated by inoculating the wild type and herbicide resistant mutant strains of A. variabilis in the presence and absence of recommended field dosages of test herbicides. No difference in survival and biofertilizer potentials of the herbicide resistant strains was observed in herbicide treated or in untreated conditions. Highest survivability (87%) was exhibited by MHR relative to other mutants. Highest growth and grain yield (76%) were recorded in plants treated with MHR as compared to uninoculated control rice plants. In conclusion, the mutant strains of A. variabilis had stable resistance to herbicides under outdoor conditions in flooded soils. Not only did the herbicide resistance strains increase growth of rice relative to the uninoculated pots, they were more beneficial for rice growth than the wild type strain. Responsible Editor: Richard W. Bell.     

Cadmium-mediated resistance to metals and antibiotics in a cyanobacterium

Summary Cadmium-resistant strains of the cyanobacterium Nostoc calcicola were isolated through the step-wise transfer of the organism to higher levels of the metal. One of the Cd-resistant strains (Cd^r–10) showed cross-resistance to antibiotics like neomycin (1 g/ml), chloramphenicol (3 g/ml) but not to streptomycin. The Cd-resistant strain also tolerated elevated levels of metals such as zinc (20 ppm) and mercury (1 ppm). The stability of the metal-resistance required the presence of Cd²⁺ ions in the growth medium. It is suggested that metal resistance may also be determined by gene(s) on the antibiotic resistance plasmids in cyanobacteria.     

蓝藻化感抑制作用研究进展

在蓝藻水华的众多危害中,对其他生物的化感抑制效应是一个重要的方面,相关研究也越来越受到生态学及环境科学领域研究者的广泛关注.系统阐述了蓝藻化感抑制作用的现象、机理及条件,总结和归纳了具有化感抑制效应的蓝藻种类、化感物质及其作用对象,讨论了蓝藻化感抑制作用的生理生态机制、影响因素、研究方法及应用前景.在此基础上提出了该领域尚未解决的问题及今后的研究方向.     

Cyanobacteria from benthic mats of Antarctic lakes as a source of new bioactivities

Aims:  To exploit the cyanobacterial diversity of microbial mats growing in the benthic environment of Antarctic lakes for the discovery of novel antibiotic and antitumour activities.
Methods and results:  In all, 51 Antarctic cyanobacteria isolated from benthic mats were cultivated in the laboratory by optimizing temperature, irradiance and mixing. Productivity was generally very low (≤60 mg l⁻¹ d⁻¹) with growth rates ( μ ) in the range of 0·02–0·44 d⁻¹. Growth rates were limited by photosensitivity, sensitivity to air bubbling, polysaccharide production or cell aggregation. Despite this, 126 extracts were prepared from 48 strains and screened for antimicrobial and cytotoxic activities. Seventeen cyanobacteria showed antimicrobial activity (against the Gram-positive Staphylococcus aureus , the filamentous fungus Aspergillus fumigatus or the yeast Cryptococcus neoformans ), and 25 were cytotoxic. The bioactivities were not in accordance with the phylogenetic grouping, but rather strain-specific. One active strain was cultivated in a 10-l photobioreactor.
Conclusions:  Isolation and mass cultivation of Antarctic cyanobacteria and LC-MS (liquid chromatography/mass spectrometry) fractionation of extracts from a subset of those strains (hits) that exhibited relatively potent antibacterial and/or antifungal activities, evidenced a chemical novelty worthy of further investigation.
Significance and impact of the study:  Development of isolation, cultivation and screening methods for Antarctic cyanobacteria has led to the discovery of strains endowed with interesting antimicrobial and antitumour activities.     

Genetic analysis of two new mutations resulting in herbicide resistance in the cyanobacterium Synechcoccus sp. PCC 7002

Two herbicide-resistant strains of the cyanobacterium Synechococcus sp. PCC 7002 are compared to the wild-type with respect to the DNA changes which result in herbicide resstance. The mutations have previously been mapped to a region of the cyanobacterial genome which encodes oneof three copies of psbA, the gene which encodes the 32 kDa Q_b-binding protein also known as D1 (Buzby et al. 1987). The DNA sequence of the wild-type gene was first determined and used as a comparison to that of the mutant alleles. A point mutation at codon 211 in the psbA1 coding locus (TTC) to TCC) results in an amino acid change from phenylalanine to serine in the D1 protein. This mutation confers resistance to atrazine and diuron at seven times and at two times the minimal inhibitory concentration (MIC) for the wild-type, respectively. A mutation at codon 211 resulting in herbicide resistance has not previously been described in the literature. A second point mutation at codon 219 in the psbA1 coding locus (GTA to ATA) results in an amino acid change from valine to isoleucine in the D1 protein. This mutation confers resistance to diuron and atrazine at ten times and at two times the MIC for the wild-type, respectively. An identical codon change conferring similar herbicide resistance patterns has previously been described in Chlamydomonas reinhardtii. The atrazine-resistance phenotype in Synechococcus sp. PCC 7002 was shown to be dominant by plasmid segregation analysis.Abbreviations At ^r atrazine resistance - Du ^r diuron resistance - Km ^r kanamycin resistance - Ap ^r ampicillin resistance - MIC Minimum inhibitory concentration     

Role of signal peptides in targeting of proteins in cyanobacteria.

Proteins of cyanobacteria may be transported across one of two membrane systems: the typical eubacterial cell envelope (consisting of an inner membrane, periplasmic space, and an outer membrane) and the photosynthetic thylakoids. To investigate the role of signal peptides in targeting in cyanobacteria, Synechococcus sp. strain PCC 7942 was transformed with vectors carrying the chloramphenicol acetyltransferase reporter gene fused to coding sequences for one of four different signal peptides. These included signal peptides of two proteins of periplasmic space origin (one from Escherichia coli and the other from Synechococcus sp. strain PCC 7942) and two other signal peptides of proteins located in the thylakoid lumen (one from a cyanobacterium and the other from a higher plant). The location of the gene fusion products expressed in Synechococcus sp. strain PCC 7942 was determined by a chloramphenicol acetyltransferase enzyme-linked immunosorbent assay of subcellular fractions. The distribution pattern for gene fusions with periplasmic signal peptides was different from that of gene fusions with thylakoid lumen signal peptides. Primary sequence analysis revealed conserved features in the thylakoid lumen signal peptides that were absent from the periplasmic signal peptides. These results suggest the importance of the signal peptide in protein targeting in cyanobacteria and point to the presence of signal peptide features conserved between chloroplasts and cyanobacteria for targeting of proteins to the thylakoid lumen.     

Rapid mineralization of the phenylurea herbicide diuron by Variovorax sp. strain SRS16 in pure culture and within a two-member consortium

The phenylurea herbicide diuron [N-(3,4-dichlorophenyl)-N,N-dimethylurea] is widely used in a broad range of herbicide formulations, and consequently, it is frequently detected as a major water contaminant in areas where there is extensive use. We constructed a linuron [N-(3,4-dichlorophenyl)-N-methoxy-N-methylurea]- and diuron-mineralizing two-member consortium by combining the cooperative degradation capacities of the diuron-degrading organism Arthrobacter globiformis strain D47 and the linuron-mineralizing organism Variovorax sp. strain SRS16. Neither of the strains mineralized diuron alone in a mineral medium, but combined, the two strains mineralized 31 to 62% of the added [ring-U-(14)C]diuron to (14)CO(2), depending on the initial diuron concentration and the cultivation conditions. The constructed consortium was used to initiate the degradation and mineralization of diuron in soil without natural attenuation potential. This approach led to the unexpected finding that Variovorax sp. strain SRS16 was able to mineralize diuron in a pure culture when it was supplemented with appropriate growth substrates, making this strain the first known bacterium capable of mineralizing diuron and representatives of both the N,N-dimethyl- and N-methoxy-N-methyl-substituted phenylurea herbicides. The ability of the coculture to mineralize microgram-per-liter levels of diuron was compared to the ability of strain SRS16 alone, which revealed the greater extent of mineralization by the two-member consortium (31 to 33% of the added [ring-U-(14)C]diuron was mineralized to (14)CO(2) when 15.5 to 38.9 mug liter(-1) diuron was used). These results suggest that the consortium consisting of strains SRS16 and D47 could be a promising candidate for remediation of soil and water contaminated with diuron and linuron and their shared metabolite 3,4-dichloroaniline.