Vol. 57, No. 1 APPLIED AND ENVIRONMENTAL MICROBIOLOGY, Jan. 1991, p. 19-28 0099-2240/91/010019-10$02.00/0 Copyright © 1991, American Society for Microbiology Influence of the Size of Indigenous Rhizobial Populations on Establishment and Symbiotic Performance of Introduced Rhizobia on Field-Grown Legumest JANICE E. THIES, PAUL W. SINGLETON, AND B. BEN BOHLOOL* University of Hawaii, NifTAL Project, 1000 Holomua Avenue, Paia, Hawaii 96779-9744 Received 1 August 1990/Accepted 14 October 1990 Indigenous rhizobia in soil present a competition barrier to the establishment of inoculant strains, possibly leading to inoculation failure. In this study, we used the natural diversity of rhizobial species and numbers in our fields to define, in quantitative terms, the relationship between indigenous rhizobial populations and inoculation response. Eight standardized inoculation trials were conducted at five well-characterized field sites on the island of Maui, Hawaii. Soil rhizobial populations ranged from 0 to over 3.5 x 104 g of soil-1 for the different legumes used. At each site, no less than four but as many as seven legume species were planted from among the following: soybean (Glycine max), lima bean (Phaseolus lunatus), cowpea (Vigna unguiculata), bush bean (Phaseolus vulgaris), peanut (Arachis hypogaea), Leucaena leucocephala, tinga pea (Lathyrus tingeatus), alfalfa (Medicago sativa), and clover (Trifolium repens). Each legume was (i) inoculated with an equal mixture of three effective strains of homologous rhizobia, (ii) fertilized at high rates with urea, or (iii) left uninoculated. For soybeans, a nonnodulating isoline was used in all trials as the rhizobia-negative control. Inoculation increased economic yield for 22 of the 29 (76%) legume species-site combinations. While the yield increase was greater than 100 kg ha-' in all cases, in only 11 (38%) of the species-site combinations was the increase statistically significant (P < 0.05). On average, inoculation increased yield by 62%. Soybean (G. max) responded to inoculation most frequently, while cowpea (V. unguiculata) failed to respond in all trials. Inoculation responses in the other legumes were site dependent. The response to inoculation and the competitive success of inoculant rhizobia were inversely related to numbers of indigenous rhizobia. As few as 50 rhizobia g of soil-' eliminated inoculation response. When fewer than 10 indigenous rhizobia g of soil-' were present, economic yield was significantly increased 85% of the time. Yield was significantly increased in only 6% of the observations when numbers of indigenous rhizobia were greater than 10 cells g of soil-'. A significant response to N application, significant increases in nodule parameters, and greater than 50% nodule occupancy by inoculant rhizobia did not necessarily coincide with significant inoculation responses. No less than a doubling of nodule mass and 66% nodule occupancy by inoculant rhizobia were required to significantly increase the yield of inoculated crops over that of uninoculated crops. However, lack of an inoculation response was common even when inoculum strains occupied the majority of nodules. In these trials, the symbiotic yield of crops was, on average, only 88% of the maximum yield potential, as defined by the fertilizer N treatment. The difference between the yield of N-fertilized crops and that of N2-fixing crops indicates a potential for improving inoculation technology, the N2 fixation capacity of rhizobial strains, and the efficiency of symbiosis. In this study, we show that the probability of enhancing yield with existing inoculation technology decreases dramatically with increasing numbers of indigenous rhizobia. Inoculation of legumes with exotic strains of rhizobia is a common agricultural practice intended to promote nitrogen fixation and increase crop yield. Despite improvements in inoculation methods (3, 13, 31, 34) and selection of rhizobial strains for increased nitrogen fixation capacity (16), compet- itive ability (1), and ability to withstand environmental stress (15, 17, 19), inoculation does not always lead to increased plant growth and crop yield. Plant response to inoculation is determined by a variety of factors. The presence and quality of indigenous rhizobial populations (3, 6, 11, 27), soil nitrogen (N) availability (9, 32), soil physicochemical constraints (12, 24), and climatic conditions (4) all significantly influence the ability to achieve increased crop yield through inoculation. Population density, effectiveness, and competitive ability are the primary characteristics of indigenous rhizobial pop- * Corresponding author. t Journal series no. 3494 of the Hawaii Institute of Tropical Agriculture and Human Resources. ulations that affect inoculation responses. In greenhouse studies, Singleton and Tavares (27) demonstrated that sta- tistically significant inoculation responses can be eliminated when there are as few as 20 indigenous rhizobia g of soil-' as long as the population contains some effective strains. Strains within populations of rhizobia differ significantly in their ability to supply the host plant with fixed N (effective- ness) under greenhouse conditions (24, 26, 27). Differences in the effectiveness of inoculant strains can also be demon- strated under field conditions as long as the soil is free of indigenous rhizobia (10). In the presence of an indigenous population, however, improved crop yield through inocula- tion with more effective inoculant strains is difficult to demonstrate (6, 11, 18). Successful competition for nodule sites by indigenous rhizobia is one reason for the failure to achieve a response to inoculation with elite rhizobial strains (18, 36). Both pot experiments (2) and field trials (36) demonstrated that to achieve nodule occupancy of greater than 50%, inoculant rhizobia must be applied at a rate at least 1,000 times greater 19