Symbiotic efficiency and diversity of native rhizobia isolated from climbing beans (phaseolus vulgaris l.) in Embu and Tharaka Nithi Counties, Kenya

Kenya is the eighth highest producer of common beans (Phaseolus vulgaris L.) worldwide with a production of 529,265 tons annually. Climbing beans are highly productive, and can produce yields 2 to 3 times higher than bush beans and is therefore a promising option to increase bean yields in Kenya. However, climbing bean production is greatly affected by the decline in soil fertility, particularly low nitrogen levels in the soil. Replenishment of soil nitrogen using effective rhizobia inoculants presents a sustainable solution to climbing bean production. The objectives of this study were to assess climbing bean varieties grown by smallholder farmers in Eastern Kenya, to determine the genetic diversity of native rhizobia strains that nodulate climbing beans and to determine nodulation and symbiotic nitrogen fixation efficiencies of native rhizobia strains isolated from climbing beans grown in Eastern Kenya. Questionnaires were used to obtain information on climbing bean varieties grown in Eastern Kenya. The experiments involved laboratory analysis, greenhouse and on farm activities. A complete randomized design and a randomized complete block design were used in the greenhouse and field experiments respectively. Five climbing bean varieties were identified in Embu and Tharaka Nithi Counties of Eastern Kenya; Gatune, Raila, Mama safi, Kithiga and Muviki, of which Gatune was the most preferred variety. In regards to bean diversity, Embu Lower Midland zone had the highest Shannon-Wiener diversity index (H’) of 2.01, while Embu Upper Midland zones had the lowest diversity index (H’) of 1.83. From the laboratory analysis, 41 native rhizobia isolates were isolated from the root nodules of mid-altitude climbing beans (MAC 13 and MAC 64) and placed into 9 groups based on their morphological characteristics. Rhizobia diversity was determined based on restriction digestion of PCR amplified 16S rRNA genes using Msp I, EcoR I and Hae III enzymes. Analysis of molecular variance based on restriction digestion of 16S rRNA genes showed that there was a 100 % genetic variation within population and no variation (0 %) among the four rhizobia populations and across the two regions (0 %). Nine representative native rhizobia isolates were tested in the greenhouse for their symbiotic nitrogen fixation efficiency on MAC 13 and MAC 64 climbing bean varieties. Four native isolates ELM3, ELM5, ELM8 and ELM4 with symbiotic efficiency (SE) of 123.72%, 99.21%, 98.24% and 96.75%, respectively, compared favorably with the commercial rhizobia inoculant (Biofix-CIAT 899) (SE of 95.21%). The best native rhizobia isolate (ELM3) was evaluated in the field experiment using MAC 13 and MAC 64 climbing beans. The mean nodule number of climbing beans differed significantly (p < 0.0001) among the test isolates. Climbing beans inoculated with test isolate ELM3 recorded a higher mean nodule number (85.58 plant-1) compared to the Biofix-CIAT 899 (76.13 plant-1) and non-inoculated control plants (52.08 plant-1). There was a significant effect of rhizobia isolates on shoot dry weight (p < 0.0001) and total bean seed yield (p < 0.0001). Climbing beans inoculated with native isolate ELM3 produced the highest mean seed yield of 4,397.75 kg/ha, indicating 89 % increase over non-inoculated control which yielded 2,334.81 kg/ha. This study demonstrated the presence of diverse native rhizobia isolates that are potentially superior over the commercial inoculant (Biofix-CIAT 899) bean inoculants. However, these isolates need to be identified and tested further in different geographical locations to determine their efficiency and stability on bean production.
A thesis submitted in partial fulfilment of the requirements for the award of the degree of Master of Science (microbiology) in the school of pure and applied sciences of Kenyatta University November, 2016