Journal of Bioscience and Biotechnology Discovery Volume 4(4), pages 60-68, August 2019 Article Number: F3A0E3081 ISSN: 2536-7064 https://doi.org/10.31248/JBBD2019.100 https://integrityresjournals.org/journal/JBBD Full Length Research Morphological and physiological characterization of nitrogen fixing rhizobia isolated from country bean (Lablaba perpureus) of Narail, Bangladesh Syed Raju Ali 1* , A. S. M. Mahadiuzzaman 1 , Rozely Hossain 1 , Suraia Nusrin 1 and Sabina Yasmin 2 1 Department of Genetic Engineering and Biotechnology, East West University, Dhaka, Bangladesh. 2 Department of Biotechnology, Bangladesh Agricultural University, Mymensingh, Bangladesh. *Corresponding author. Email: syedrajuali@gmail.com Copyright © 2019 Ali et al. This article remains permanently open access under the terms of the Creative Commons Attribution License 4.0, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. Received 18th June, 2019; Accepted 14th August, 2019 ABSTRACT: Nitrogen is one of the important constituents of the plants. It is fixed in legume plants from the atmosphere by the association of beneficial gram negative soil bacteria named rhizobia. This study aimed to distinguish the morpho- physiological characteristics of naturally growing rhizobia having higher environmental and external stress tolerance. Twenty-two isolates were isolated from country bean root nodules and characterized based on different tests. The isolates varied from one another with level of parameters of the treatments. Most of them exhibited cream color colonies and some isolates evolved white and pink colonies. The average colony size of the isolates was ranged between 0.83 to 2.83 mm after one day and 1.33 to 5.00 mm after two days. Among them, 17 isolates were alkali producers, three isolates were acid producer and only two isolates produced neutral mucous. All the isolates showed complete resistance at pH 5.0 to 10.0, 2% NaCl, 10 to 42 o C temperature, and 0.1 mM of heavy metallic salt MnCl2, FeCl3 and PbCl3. Nevertheless, most of the isolates also tolerated pH 4.0; 3 to 7% NaCl; 47 o C temperature; 0.1 mM CuSO4. On the other hand, a few numbers of the isolates resisted 100 μg/ml and 200 μg/ml ampicillin, kanamycin and tetracycline antibiotics. Therefore, exploration of country bean rhizobia for their environmental stress tolerance capacity might be a core study for the production of cost- effective bio-fertilizer to boost up country bean yield. Keywords: Country bean, Nitrogen fixation, nodule, rhizobia, symbiosis. INTRODUCTION The biological fixation of atmospheric nitrogen is carried out by a symbiotic relationship between the plant and bacteria. It is one of the main sources of naturally nitrogen pool enhancement in cultivation lands. Rhizobia, a group of useful gram negative soil bacteria, induce nodulation on leguminous plants and perform nitrogen fixation through symbiosis process (Zhao-Hai et al., 2007). The bacteria contain some nitrogenase enzymes which reduces atmospheric free nitrogen to ammonia or other nitrogenous complexes. Rhizobia consume required carbon source from the host plant and supply the nitrogenous compounds to the plants. Such type of successful symbiotic association helps rhizobia to survive and promote the adequate number in the soil ecosystem (KÜÇÜK, 2011). The higher the survival amount of rhizobium in leguminous nodule, the greater the yielding rates of legume product. Lablaba perpureus is a leguminous species native to Asia and Africa, used in Bangladesh as commercially cultivating winter vegetable, known as 'Sheem' (Pengelly and Maass, 2001; Saha and Haque, 2005). As L. perpureus contains high amount of protein, it can be harvested to reduce the protein scarcity existing in the developing and underdeveloped countries (Adebowale and Lawal, 2003). The high productivity of L. perpureus can be done by increasing their natural biological nitrogen fixation (BNF) capability. To enhance this biological nitrogen fixation (BNF) potential of this crop, the selection