Screening for Symbiotically Effective and Ecologically Competitive Chickpea Rhizobial Inoculants from Ethiopian Soils
Abstract
Chickpea is one the most important pulse crops grown for food and crop rotation in order to improve soil fertility because it fixes atmospheric nitrogen with root nodule bacteria. However, effectiveness in nitrogen fixation depends upon the host variety, the efficient endosymbiont and environmental conditions. This study was initiated to isolate and characterize chickpea rhizobia for their symbiotic effectiveness adapted to local environmental conditions. A total of seventy root nodule bacteria were isolated from different sampling sites in central and northern Ethiopia of which only 52% were rhizobia and the remaining were endophytes and non-nodulating rhizobia. All but two of the isolates were fast growers with generation time (<3 h) with large mucoid and large watery colony texture, and colony size (1-5 mm) and were able to change BTB-YEMA medium to yellow indicating that they were fast growing Metarhizobium ciceri. The isolates utilized different carbohydrates, except dulcitol, aesculine and citrate, and many amino acids, but none of the isolates assimilated aspargine and arginine, and a few isolates were able to utilize glycine and thymine. The isolates showed a pattern of tolerance to high pH (>8.0), high salt (4-6%) and medium temperature (20-30°C), and were sensitive to low pH (pH 4-4.5) and high temperature (40-45°C). Most isolates were resistant to several antibiotics and heavy metals, but relatively sensitive to chloramphenicol, tetracycline (51%). Of the tested antibiotics and heavy metals, streptomycin sulfate and mercury were found to be the most potent against the isolates. The symbiotic effectiveness test showed that only 20% of the isolates were able to increase the shoot dry mass as much as 50-100%. Of the nitrogen-fertilized control plants, two isolates (isolates NSCPR13 and NSCPR14) were highly effective (80%-100%). Based on their symbiotic effectiveness and wide physiological diversity and tolerance, the two best isolates; NSCPR13 and NSCPR24 could be tested in the field as future candidates for commercial inoculation.
Downloads
References
Alikhani, H.A., Saleh-Rastin, N. and Antoun, H. (2006). Phosphate solubilization activity of Rhizobia native to Iranian soils. Plant Soil. 287: 35–41.
Amarger, N., Macheret, V. and Laguerre, G. (1997). Rhizobium gallicum spp. Nov. and Rhizobium giardinii spp. Nov. from Phaseolus vulgaris nodules. Int. Syst. Bacteriol.47: 996–1006.
Angaw Tsigie and Asnakew Woldeab (1994). Fertilizer response trials on highland food legumes of Ethiopia. In: Cool Season Food Legumes of Ethiopia, pp. 279–292 (Asfaw Tilaye, Geletu Bejiga, Saxena, M.C. and Solh, M.B., eds.). International Centre for Agricultural Research in the Dry Areas, Addis Ababa, Ethiopia.
Asfaw Tilaye, Geletu Bejiga and Alem Berhane (1994). Role of cool season food legumes of Ethiopia. In: Cool Season Food Legumes of Ethiopia, pp. 3–19 (Asfaw Tilaye, Geletu Bejiga, Saxena, M.C. and Solh, M.B., eds.). International Centre for Agricultural Research in the Dry Areas, Addis Ababa, Ethiopia.
Beck, D.P. (1992). Yield and nitrogen fixation of chickpea cultivars in response to inoculation with selected Rhizobia strains. Agron. J. 84: 510–516.
Broughton, W.J. and Dilworth, M.J. (1971). Control of leghaemoglobin synthesis in snake beans. Biochem. J. 125: 1075–1080.
CSA (Central Statistical Authority) (2008). Agricultural sample survey 2007/2008. Report on area and production of crops. Statistical bulletin volume I, number 417, Addis Ababa, Ethiopia.
Danso, S.K., Quander, M.A. and Sattar, M.A. (1995). Nodulation, nitrogen fixation and yield of chickpea as influenced by host cultivar and Bradyrhizobium strain difference. Soil Biol. Biochem. 27: 725–727.
Date, R.A. (1993). Assessment of Rhizobial status of the soil. In: Nitrogen Fixation in Legumes, pp. 85–94 (Vincent, J.M., ed.). Academic Press, Sydney.
El Hadi, E.A. and Elsheikh, E.A.E. (1999). Effect of Rhizobium inoculation and nitrogen fertilization on yield and protein content of six chickpea (Cicer arietinum L.) cultivars in marginal soils under irrigation. Nutr. Cycl. Agroecosys. 54: 57–63.
Elsheikh, E.A.E. (1992). Effect of salinity on growth, nodulation and nitrogen yield of inoculated and nitrogen fertilized chickpea (Cicer arietinum L.). Arch Biotechnol.1: 17–28.
EEPA (Ethiopian Export Promotion Agency) (2004). http://www.ethioconsulate- la.org.
Geletu Bejiga, Million Eshete and Yadeta, A. (1996). Improved cultivars and production technology of chickpea in Ethiopia. Research Bulletin No. 2. Debre-Zeit, Ethiopia, Debre-Zeit Agricultural Research Centre, Alemaya University of Agriculture.
Gemechu Keneni, Endashaw Bekele, Fassil Assefa, Muhammad Imtiaz, Tolessa Debele, Kifle Dagne and Emana Getu (2012). Phenotypic diversity for symbio- agronomic characters in Ethiopian chickpea (Cicer arietinum L.) germplasm. Afr. J. Biotechnol. 11(63): 12634–12651.
Giller, K.E. (2001). Nitrogen Fixation in Tropical Cropping Systems. 2nd ed. CABI Publishing, Walling Ford.
Graham, P.H. (1992). Stress tolerance in Rhizobium and Bradyrhizobium and nodulation under adverse soil conditions. Can. J. Microbiol. 38: 475–484.
Gregorson, G. (1978). Rapid method for distinction of Gram-positive bacteria. Eur. J. Appl. Microbiol. 5: 123–127.
Halder, A.K., Mishra, A.K. and Chakrabartty, P.K. (1990). Solubilization of phosphatic compounds by Rhizobium. Indian J. Microbiol. 30: 311–314.
Hungaria, M., Andrade, D.S. and Chueira, L.M. (2000). Isolation and characterization of new efficient and competitive bean (Phaseolus vulgaris L.) rhizobia in Brazil. Soil Biol. Biochem. 32: 1515–1528.
Jordan, D.C. (1984). Family III Rhizobiaceae. In: Bergey’s Manual of Systematic Bacteriology, pp. 234 –256 (Kreig, N.R. and Holt, J.G., eds.). Vol. 1. Williams and Williams Co., Baltimore.
Küçük, C. and Kivanç, M. (2008). Preliminary characterization of Rhizobium strains isolated from chickpea nodules. Afr. J. Biotechnol.7: 772–775.
L’taief, B., Sifi, B., Gtari, M., Zaman-Allah, M. and Lachaal, M. (2007). Phenotypic and molecular characterization of chickpea rhizobia isolated from different areas of Tunisia. Can. J. Microbiol. 53: 427–434.
Maatallah, J., Berrah, E.B., Sanjuan, J. and Lunch, C. (2002). Phenotypic characterization of Rhizobia isolated from chickpea (Cicer arietinum) growing in Moroccan soils. Agronomie 22: 321–329.
Martha, L., Machado, J., Young, W. and Oliveira, S. (2004). High diversity of chickpea-Mesorhizobium species isolated in a Portuguese agricultural region. FEMS Microbiol. Ecol. 48: 101–107.
Mahdi, A.A. (1992). The biofertilizer use of Rhizobium strain TAL 634 and CM 127 for bean and chickpea in the Sudan. Arch Biotechnol.1: 10–16.
Mulissa Jida and Fassil Assefa (2012). Phenotypic diversity and plant growth promoting characteristics of Mesorhizobium species isolated from chickpea (Cicer arietinum L.) growing areas of Ethiopia. Afr. J. Biotechnol.11: 7483–7493.
Nour, S.M., Cleyet-Marel, J.C., Beck, D., Effoss, A. and Fernando, M.P. (1994). Genotypic and phenotypic diversity of Rhizobium isolated from chickpea (Cicer arietinum L.). Can. J. Microbiol. 40: 345–354.
O’Hara, G.W., Yates, R. and Hawieson, J. (2002). Selection of strains of root nodule bacteria to improve inoculants performance and increase legume productivity in stressful environment. In: Inoculants of Nitrogen Fixation of Legumes in Vietnam, pp. 75–80 (Herridge, D., ed.). ACIAR Proceedings 109, Australia.
Peix, A., Rivas-Boyero, A.A., Mateos, P.F., Rodriguez-Barrueco, C., Martinez-Molin, E. and Velazquez, E. (2001). Growth promotion of chickpea and barley by phosphate solublizing strain of Mesorizobium mediteraneum under growth chamber condition. Soil Biol. Biochem. 33: 103–110.
Peoples, M.B., Ladha, J.K. and Herridge, D.R. (1995). Enhancing legume N2 fixation through plant and soil management. Plant Soil 174(1/2): 83–101.
Romdhane, S.B., Aouani, M.E., Trabelsi, M., De Lajudie, P. and Mhamdi, R. (2009). Selection of high Nitrogen-fixing Rhizobia nodulating chickpea (Cicer arietinum) for semi-arid Tunisia. J. Agron. Crop. Sci. 194: 413–420.
Sahlemedhin Sertsu and Taye Bekele (2000). Procedures for Soil and Plant Analysis. National Soil Reseach Centre. EARO, Ethiopia.
Sharma, M.P., Srivatava, K. and Sharma, S.K. (2010). Biochemical characterization and metabolic diversity of soybean rhizobia isolated from Malwa Region of central India. Plant Soil Environ. 56: 375–383.
Somasegaran, P. and Hoben, H.J. (1994). Handbook for Rhizobia: Methods in Legume-Rhizobium Technology. Springer, Berlin, Heidelberg, New York.
Vincent, J.M. (1970). A Manual for the Practical Study of Root Nodule Bacteria. Blackwell, Oxford and Edinburgh.
Werner, D. (2005). Production and biological nitrogen fixation of tropical legumes. In: Nitrogen Fixation in Agriculture, Forestry, Ecology, and the Environment, pp. 1–13 (Werner, D. and Newton, W.E., eds.). Springer, Netherlands.
White, D. (1995). The Physiology and Biochemistry of Prokaryotes. Oxford University Press, New York.
Zohary, D. and Hopf, M. (1993). Domestication of Plants in the old World – The Origin and Spread of Cultivated Plants in West Asia, Europe, and the Nile Valley. Clarendon Press, Oxford.
