EFFECT OF INOCULATION DENSITY OF AZOLLA FRONDS ON NITROGENASE ACTIVITY AND HETEROCYST FREQUENCY OF THE AZOLLA-ANABAENA AZOLLAE SYMBIOSIS

Document Type : Original Article

Authors

1 Botany Department, Faculty of Science, Cairo University, Giza, Egypt

2 Agricultural Microbiology Dept., Soils, Water & Environ. Res. Inst., Agric. Res. Center, Giza, Egypt

Abstract

The effect of inoculation density level for three Azolla species namely A. microphylla, A. filiculoides and A. pinnata was investigated under greenhouse conditions in term of senescence associated with Azolla fronds overcrowding. In this respect three inoculum levels, i.e., 0.2 kg, 1.0 kg and 2.0 kg m-2 were tested.
 Overcrowding influence on Azolla was assessed in terms of frond yellowing, nitrogenase activity and heterocyst frequency. Frond discoloration of A. microphylla, A. filiculoides was observed after only 7 days in higher inocula level (1.0 and 2.0 kg m-2). However, in the lower level of inocula 0.2 kg m-2 the discoloration was observed after 15 days. However, A. pinnata exhibited frond discoloration on day 10 for 0.2 kg and on day 5 for higher inoculum levels. The nitrogenase activity increased significantly on 7 days at less crowded condition in the cultures inoculated with 0.2 kg m-2 for all the tested species. At higher inoculum levels, the nitrogenase activity declined on the third week, after which very low enzyme activity, overcrowding, and extensive senescence were noticed at all inoculurn levels. Azolla plants maintained in the dark for 12 hrs prior to beginning of dark incubation period of 3 hrs showed an almost zero nitrogen-fixing activity compared to plants exposed to continuous light. Heterocyst formation on the 12th leaf from the tip increased up to the seventh day in the three species at 0.2 kg m-2and in A. pinnata only at 1.0 kg m-2 inoculum level. There was an apparent gradually decline in the heterocyst count in the three species at higher inoculum levels throughout the culture period.

Ashton, P. J. and Walmsley. (1976). The aquatic fern Azolla and its Anabaena symbiont. Endeavor, 35:39-47.
Bar, E.; Kulasooriya, A. and Tel-Or, E. (1991). Regulation of nitrogenase activity in the Azolla-anabaena symbiosis. Bioresource Technology, 38: (2 and 3), 171-177.
Bar, E. and Tel-Or, E. (1994). Effect of light and oxygen on nitrogenase and dinitrogenase reductase (Fe-protein) content in Azolla-anabaena association. Journal of Plant Physiol., 144: (4-5): 438-443.                  
Becking, J. H. (1976). Contribution of plant-algal association In WE Newton, CJ Nyman, eds, Proceedings of the first international symposium on nitrogen fixation. Vol.2. Washington State University Press, Pullman pp. 556-558.
Cohn, M. and Thimann, K. (1972). The role of protein synthesis in the senescence of leaves. 1- The formation of protease. Plant Physiol., 49: 64-71.
Chon, H. T. and Thimann, K. (1975). The metabolism of oat leaves during senescence. 3- The senescence of isolated chloroplast. Plant Physiol., 55: 828-834.
Fogg, G. E.; Stewart, W. D. P.; Fay, P. and Walsby, A. E. (1973). The blue-green algae. 235, (Academic Press, London, New York).
Ghazal, F. M. and Herzalla, Nagat A. (1997). Effect of Senescence of Azolla fronds on nitrogenase activity and heterocyst frequency of the Azolla-Anabaena Azolla Symbioses. Al-Azhar J Agric. Res., 25, 98 – 114.
Gomez, K. A. and Gomez, A. A. (1984). Statistical Procedures For Agricultural Research. 2nd ed. pp. 20-29 & 3 59-487.
Hardy, R.F.; Burns R. C. and Hoisten, R. D. (1973). Application of the acetlylene-ethylene assay for measurement of nitrogen - fixation. Soil Biol. Biochem., 5:47-81.
Hill, D. J. (1975). The pattern of development of Anabaena in the Azolla-Anabaena symbiosis. Planta, 122: 179-184.
International Rice Research Institute (IRRI) (1985). Nitrogen Fixation and Azolla Management. Annual Report, P. 340-253.
Lex, M. and Stewart, W. D. P. (1973). Algal nitrogenase reductant pools and photosystem I. activity. Biochiem. Biophys. Acta., 292: 436-443.
Moore, A.W. (1969). Azolla: Biology and agronomic significance.Bot. Rev. 35 17-35. Munchen, F.R., Sheeny, J.E. and Witty, J.F.(1986). Further errorsin the acetylene reduction assay. Effect of plant disturbance. J. Expt. Bot., 37: 1581-1591.
Peters, G. A. (1975). Studies on the Azolla-Anabaena Azollae relationship. III. Studies on metabolic capabilities and further characterization of the symbiont. Arch. Micrbiol., 103: 112-113.
Peters, G. A. (1976). Studies on the Azolla-Anabaena Azollae symbiosis. In WE Newton, CJ Nyman, eds, Proceedings of the first international sympsoium on nitrogen fixation. Vol. 2. Washington State University Press, Pullman pp. 295-6 10.
Peters, G. A. and Mayne, B. C. (1974). Azolla-Anabaena Azollae relationship. I-Initial characterization on the association. Plant Physiol., 53, 813-819.
Peters, G. A.; Evans, W. R and Toia, J. R. (1976). Azolla-Anabaena Azollae relationship. IV-Photosynthetically driven, nitrogenase catalyzed H2 production. Plant Physiol., 58: 118-126.
Peters, G. A.; Robert, E.; Toia, J. R. and Samuel, M. L. (1977). Azolla-Anabaena Azollae relationship. V- 15N2 Fixation, Acetylene reduction, and H2 production. Plant Physiol., 59: 1021- 1025.
Tung, H. F. and Watanabe, L. (1983). Accumulation of ammonium-N and Amino-N in the Azolla-Anabaena  association. Plant and Soil, 73: 413-419.
Uheda, F. (1986). Isolation of empty packet from Anabaena-free Azolla. Plant Physiol., 81: 1187-1 190.
Yoshida, S.; Forno, D. A.; Cock, J. H. and Gomez, K.A. (1976). Laboratory Manual for Physiological studies of rice. The International Rice Research Institute, Manila, Philippines.
Vaishampayan, A. (1982). Cu-Fe interaction in the N2 -fixing cyano bacterium Nostoc muscorum. Microbios Letters, 21: 17-23.
Watanabe, I. and Berja, S. (1983). The growth of four species of Azolla as affected by temperature. Aquatic Botany, 15: 175-185.