Mayashree B. Syiem

Zn(II) and Cu(II) removal by Nostoc muscorum: a cyanobacterium isolated from a coal mining pit in Chiehruphi, Meghalaya, India

Nostoc muscorum was isolated from a coal mining pit in Chiehruphi, Meghalaya, India, and its potential to remove Zn(II) and Cu(II) from media and the various biochemical alterations it undergoes during metal stress were studied. Metal uptake measured as a function of the ions removed by N. muscorum from media supplemented independently with 20 μmol/L ZnSO4 and CuSO4 established the ability of this cyanobacterium to remove 66% of Zn(2+) and 71% of Cu(2+) within 24 h of contact time. Metal binding on the cell surface was found to be the primary mode of uptake, followed by internalization. Within 7 days of contact, Zn(2+) and Cu(2+) mediated dissimilar effects on the organism. For instance, although chlorophyll a synthesis was increased by 12% in Zn(2+)-treated cells, it was reduced by 26% in Cu(2+)-treated cells. Total protein content remained unaltered in Zn(2+)-supplemented medium; however, a 15% reduction was noticed upon Cu(2+) exposure. Copper enhanced both photosynthesis and respiration by 15% and 19%, respectively; in contrast, photosynthesis was unchanged and respiration dropped by 11% upon Zn(2+) treatment. Inoculum age also influenced metal removal ability. Experiments in the presence of 3-(3,4-dichlorophenyl)-1,1-dimethylurea (a photosynthetic inhibitor), carbonyl cyanide m-chlorophenyl hydrazone (an uncoupler), and exogenous ATP established that metal uptake was energy dependent, and photosynthesis contributed significantly towards the energy pool required to mediate metal removals.

Isolation and characterization of a Mastigocladus species capable of growth, N(2)-fixation and N-assimilation at elevated temperature.

A Mastigocladus species was isolated from the hot spring of Jakrem (Meghalaya) India. Uptake and utilization of nitrate, nitrite, ammonium and amino acids (glutamine, asparagine, arginine, alanine) were studied in this cyanobacterium grown at different temperatures (25°C, 45°C). There was 2–3 fold increase in the heterocyst formation and nitrogenase activity in N-free medium at higher temperature (45°C). Growth and uptake and assimilation of various nitrogen sources were also 2–3 fold higher at 45°C indicating that it is a thermophile. The extent of induction and repression of nitrate uptake by NO3− and NH4+, respectively, differed from that of nitrite. It appeared that Mastigocladus had two independent nitrate/nitrite transport systems. Nitrate reductase and nitrite reductase activitiy was not NO3−-inducible and ammonium or amino acids caused only partial repression. Presence of various amino acids in the media partially repressed glutamine synthetase activity. Ammonium (methylammonium) and amino acid uptake showed a biphasic pattern, was energy-dependent and the induction of uptake required de novo protein synthesis. Ammonium transport was substrate (NH4+)-repressible, while the amino acid uptake was substrate inducible. When grown at 25°C, the cyanobacterium formed maximum akinetes that remained viable upto 5 years under dry conditions.

N 2 -Fixing Cyanobacterial Systems as Biofertilizer

Soil and water surfaces, as well as plant surfaces and tissues are the known locations that harbor free-living phototrophic N2-fixing cyanobacteria. These organisms are known to contribute substantial amounts of fixed nitrogen (20–30 kg N ha−1annually). In continents where rice is the prime crop for majority of the population (amounting to over 40 % of world’s population), these organisms assume great importance. Two third of the total of 180 million tons of fixed nitrogen that gets added to the earth’s surface globally, comes from biological activities mainly contributed by these and other microbes. Rice field ecosystems are ideal for cyanobacterial growth as they provide optimum growth conditions. Azolla-Anabaena symbiotic association, another cyanobacterial system has been exploited as a biofertilizer in many Asian countries. This symbiosis is very important agronomically because its contribution has been estimated to be ~600 kg N ha−1. With the adverse consequences of chemical agriculture, focus on nitrogen enrichment has shifted again to biological nitrogen fixation, especially towards both free-living and symbiotic cyanobacteria. During past few decades, research studies have yielded a large quantity of information on cyanobacterial nitrogen fixation from isolation, molecular understanding and manipulations to large-scale production for agriculture. Substantial research studies have also been devoted towards creating and understanding the artificial associations of cyanobacteria with crop plants. In this chapter, various N2-fixing cyanobacterial systems in light of their use as biofertilizers are reviewed.

A Common Transport System for Methionine, l -methionine- dl -Sulfoximine (MSX), and Phosphinothricin (PPT) in the Diazotrophic Cyanobacterium Nostoc muscorum

We present evidence, for the first time, of the occurrence of a transport system common for amino acid methionine, and methionine/glutamate analogues l-methionine-dl-sulfoximine (MSX) and phosphinothricin (PPT) in cyanobacterium Nostoc muscorum. Methionine, which is toxic to cyanobacterium, enhanced its nitrogenase activity at lower concentrations. The cyanobacterium showed a biphasic pattern of methionine uptake activity that was competitively inhibited by the amino acids alanine, isoleucine, leucine, phenylalanine, proline, valine, glutamine, and asparagine. The methionine/glutamate analogue-resistant N. muscorum strains (MSX-R and PPT-R strains) also showed methionine-resistant phenotype accompanied by a drastic decrease in 35S methionine uptake activity. Treatment of protein extracts from these mutant strains with MSX and PPT reduced biosynthetic glutamine synthetase (GS) activity only in vitro and not in vivo. This finding implicated that MSX- and PPT-R phenotypes may have arisen due to a defect in their MSX and PPT transport activity. The simultaneous decrease in methionine uptake activity and in vitro sensitivity toward MSX and PPT of GS protein in MSX- and PPT-R strains indicated that methionine, MSX, and PPT have a common transport system that is shared by other amino acids as well in N. muscorum. Such information can become useful for isolation of methionine-producing cyanobacterial strains.

Dose dependent variance in UV-C radiation induced effects on carbon and nitrogen metabolism in the cyanobacterium Nostoc muscorum Meg1

With the intention of getting an insight into the differential effect of UV-C radiation on the N2-fixing heterocystous cyanobacterium Nostoc muscorum Meg1, various aspects of carbon and nitrogen metabolism was evaluated in the organism. Exposure to different doses of UV-C (6, 12, 18 and 24 mJ/cm2) showed that among various photo-absorbing pigments, phycobiliproteins were most sensitive. Oxygen evolving complex (OEC) activity measured as net oxygen evolution rate decreased by 63% upon 24 mJ/cm2 exposure. Western blot analysis established that D1 protein of PSII was highly sensitive and its levels decreased even at a radiation dose as low as 6 mJ/cm2. In contrast, levels of the Calvin cycle enzyme RuBisCO was increased at 6 and 12 mJ/cm2 doses but the level decreased drastically (84%) at higher dose (24 mJ/cm2). The nitrogenase enzyme activity decreased at all doses but the ammonia assimilating enzyme glutamine synthetase (GS) activity recorded increase at the lower doses. The reactive oxygen species (ROS) and lipid peroxidation increased upon UV-C exposure. Transmission electron microscopic observation revealed damage to ultrastructure especially the thylakoid membrane organization, aggregation of dissolving phycobilisomes and loss of caboxysomes. Interestingly, sub-lethal radiation (6 and 12 mJ/cm2) dose exposures increased the growth rate in the organism when growth was measured over a period of 11 days after radiation exposure.

Protective Role of Ca2+ towards Cu2+ Induced Toxicity on Photosynthetic Pigments, Morphology and Ultra-Structures of the Cyanobacterium Nostoc muscorum Meg 1

Ca2+ has been reported to play a protective role in many cyanobacteria against toxic effects of various metals. However there are very few reports of Ca2+ mediated protection in Cu2+ treated cyanobacterial cells. An initial study conducted to assess the influence of Ca2+ over Cu2+ induced effects on morphology, ultra-structure, photosynthetic pigments and total protein content of cyanobacterial Nostoc muscorum Meg 1 revealed that as little as 3 ppm Cu2+ can induce reduction in all these parameters by 50-80%. However when 10 ppm Ca2+ was present along with 3 ppm Cu2+, the Cu2+ induced toxic effects were lessened by 55-85% within 7 days. Bright field and scanning electron microscopic study showed that morphological changes including broken filaments; rupture, elongation and shrivelling of cells were lessened upon inclusion of Ca2+. Ultra-structural studies conducted using transmission electron microscopy showed detachment of cell membrane from cell wall, shrinkage of cellular matter; compromised thylakoid membranes and increased number of polyphosphate bodies in the Cu2+ treated cells whereas these effects were convincingly less in presence of Ca2+. Similarly decrease in protein concentration under the influence of Cu2+ was also positively modulated by the presence of Ca2+.