Evaluation of Methods for Estimating Numbers of Rhizobia on Alfalfa Seed1

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Abstract
Comparative studies were made to determine the relative efficiency of different laboratory tests in detecting Rhizobium meliloti when alfalfa seeds were inoculated with known numbers of bacteria. The bottomless bottle (modified Leonard) and crock techniques, which utilize more than one seed per container, overestimated the number of rhizobia present on alfalfa seeds due to the transfer of rhizobia among seedlings in the test container. The transfer effect probably was due to root growth accompanied by multiplication and movement of rhizobia. Positive results (nodulated plants) in a container indicated that at least one seed carried one rhizobium. When rhizobia were present on the seed, a container with one inoculated seed gave results that were not significantly different from the results obtained when all seeds were inoculated. The agar tube technique tended to underestimate the number of rhizobia present on alfalfa seeds because the anaerobic conditions in the support medium inhibited nodule formation. The sand tube technique gave the most accurate information.

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