Rhizobium - Host Specificities in Pbaseolus coccineus L. and Pbaseolus vulgaris L

The introduction of desirable agronomic characteristics into Phaseolus vulgaris L. is being attempted by interspecific hybridization with P. coccineus, but the effects on root nodulation are unknown. Accordingly, cross-inoculation relationships between P. vulgaris `Red Kidney' and P. coccineus ’Scarlet Runner' were tested in the greenhouse with a group of 16 Rhizobium strains effective on Red Kidney and a group of 12 strains effective on Scarlet Runner. Both groups had diverse origins, including commercial sources and isolations from bean fields and other sites in California. Only six strains, all isolates from Scarlet Runner, were effective on both hosts. Fifty percent of Scarlet Runner isolates were as effective on Red Kidney as the more effective of Red Kidney's homologous strains. Thirty percent of the California isolates tested on Red Kidney were more effective than two commercial inocula. Strain rejection in Red Kidney was expressed in 75 % of all cases as failure to form nodules. In contrast, strain rejection by Scarlet Runner was expressed in 64 to 68 % of all cases as ineffective nodulation (nodule malfunction). If Scarlet Runner is representative of P. coccineus in this respect, interspecific hybridization is likely to increase rather than reduce the tendency of P. vulgaris to nodulate with ineffective rhizobia. Additional index words: Rhizobium phaseoli, Phaseolus vulgaris, Phaseolus coccineus, Cross-inoculation, Nodulation effectiveness, Isolation of soil rhizobia. MODERN cultivars of Phaseolus coccineus L. and P. vulgaris have evolved from related wild forms of the genus Phaseolus (6, 7, 10, 11, 17, 18). Cultivars of P. vulgaris exhibit a greater diversity of habit and other characteristics than do those of P. coccineus (6, 18), but the latter are thought to show greater vegetative vigor, seed size, disease resistance, and adaptation to cooler climates. These have been desirable characteristics for breeding programs aimed at improving P. vulgaris cultivars (13, 16). Graham and Halliday (8) have suggested interspecific hybridization as a means of increasing host nodulation specificity in P. vulgaris and improving symbiotic performance under field conditions. Phaseolus vulgaris has often been noted for its promiscuity (1, 8, 9, 20, 21), and for the detrimental effects of ineffective nodulation (1, 2, 12, 14, 15). Phaseolus coccineus may be resistant to nodulation by strains of Rhizobium phaseoli effective on P. vulgaris, but current information is limited (1, 8, 21). 'Contribution from Dep. of Land, Air and Water Resources and Dep. of Agronomy, Univ. of California, Davis CA 95616. Received 16 October 1980. 'Graduate research assistant and professor of Soil Science, Dep. of Land, Air and Water Resources; specialist, Dep. of Agronomy, respectively, Univ. of Calif., Davis, CA 95616. In view of these breeding interests and concern with improving N fixation in P. vulgaris, a study of nodulation characteristics in P. coccineus seems appropriate. This paper presents data on rhizobial nodulation specificities for one cultivar of each species: P. vulgaris `Red Kidney' and P. coccineus `Scarlet Runner'. Nodulation characteristics and overall effectiveness of each host were tested with commercial bean rhizobia and native strains from 16 California fields. MATERIALS AND METHODS Effectiveness of selected rhizobial strains was determined on P. coccineus `Scarlet Runner' and P. vulgaris `Red Kidney' under greenhouse conditions. Three trials were conducted in the spring and early summer of 1978 and 1979. Scarlet Runner seeds were obtained commercially from Northrup King Co. for the host isolation trial and screening Trial 1, and from Lagomarsino Seed Co. for screening Trial 2. Red Kidney seed was supplied by C. L. Tucker, Dep. of Agronomy and Range Science, Univ. of California, Davis. Eighteen California soils were sampled (Table 1). Two pots were filled with 2 kg of each soil, air-dry and unamended. Pots were maintained at approximate field capacity by watering to weight with distilled water. Two surface-sterilized seeds of each cultivar were planted in a pot. After emergence, seedlings were thinned to one plant per pot. Plants were harvested 30 days after emergence, and crushed nodule isolates were obtained from the 2 CROP SCIENCE, VOL. 22, MAY JUNE 1982 largest three proximal nodules on each plant by the procedure of Vincent (19). Uniform, single-celled colonies were cultured on yeast mannitol agar. Cultures were discarded if they were not short, gram negative rods. Forty-four strains were retained. In addition, established strains for P. vulgaris and Vigna spp. were included (Table 2). In all trials, seeds were surface sterilized by immersion in 5 % H202 for 5 min, and planted at 5 cm depths at a rate of four per pot. Seedlings were thinned for uniformity to two per pot. Treatments were replicated by four pots in Trial 1 and by three pots in Trials 2 and 3. Pots were arranged in randomized complete blocks and watered to weight with distilled water throughout growth. Plant color, budding, and flowering were noted relative to that of the uninoculated control treatments, with and without NH4NO3. After harvesting, plant tops were dried at 65 C and weighed. Nodule count, weight, color, and distribution were recorded. Acetylene reduction rate was measured during harvest between 1000 hours and 1400 hours. For this purpose, plant root systems were cut off below the cotyledon, placed in 970 ml canning jars fitted with serum stoppers, and incubated for 1 hour in a 10% carbide-generated acetylene atmosphere. Samples were then withdrawn for gas chromatography. Trial 1. Test of effective common bean rhizobia on Scarlet Runner. The growth medium was river outwash from Cache Creek, Calif., with sandy loam texture and pH 7.8. Pots with plastic liners were filled with 2 kg of dry, autoclaved soil. A basal fertilizer was mixed with the soil in each pot 1 day prior to planting, providing 3 mmole/pot of both KH2P04 and K2S04, 10 ppm Zn as ZnSO4, and 0.1 ppm Mo as Na2MoO4. Inoculum was prepared from fresh rhizobial suspensions estimated to provide 10 viable cells/ml, and 2 ml/seed were applied at planting. Nitrogen fertilized controls received 4 mmole NH4NO3 2 weeks after germination. Soil temperature fluctuated between 20 and 28 C. A 15% moisture content was maintained throughout growth. Plants were harvested 47 days after emergence. Trial 2. Test of Scarlet Runner isolates on Scarlet Runner. Pots containing a mix of 2:1 horticultural perlite and fine quartz sand were watered with 400 ml of nutrient solution containing 3.75 m M CaC12, 1.25 m M MgSO„ 1.25 m M K2S0„ 0.38 m M KH2P0„ 0.25 m M K2HP0„ 62.5,u M Fe-EDTA, and micronutrients. The growth medium was autoclaved for 2 to 3 hours. Nutrient solution was again added 19 days after germination. Inoculum was prepared from fresh rhizobial suspensions esti Fig. 1. Growth and nodulation of Scarlet Runner with rhizobia effective on P. vulgaris, P. coccineus, and Vigna spp. Strains identified by numbers according to Tables 1 and 2. Dotted line separates effective from ineffective symbioses, based on differences in plant top dry wt. (P + 0.05). mated to provide 10 viable cells/ml, and seedlings each received 1.5 ml 3 to 5 days after emergence and thinning. The N fertilized controls received 5 mmole of NH4NO3 both 15 and 29 days after germination. Soil temperatures were kept between 23 and 28 C by placing pots in constant temperature water baths. Forty percent moisture content was maintained in the pots. Plants were harvested from 34 to 36 days after emergence. Trial 3. Test of Scarlet Runner and Red Kidney isolates on Red Kidney. A combination of medium sand, 10% by volume spaghnum peat, and distilled water was autoclaved, placed in polyethylene lined pots and planted as in Trial 2. After emergence, the pot surfaces were covered with sterilized coarse perlite to reduce cross-contamination. Nutrient solution containing 1 m M CaC12, 1 m M MgSO4, 1 m M KH,2PO4, 10 μ M FE-EDTA, and micronutrients was added in increments; 0.25 strength the 1st week after germination, 0.50 strength during the 2nd week, and full strength during the remaining 3 weeks. The water soluble N content of the sand was below detectable levels ( < 0.016 mmole/ 100 g soil) and that of the peat was 1.06 mmole N/pot. The N fertilized controls received an additional 5 mmole NH4NO3 13 and 28 days after emergence. Soil temperatures fluctuated from 24 to 31 C during the day and from 17 to 21 C during the night. Pots were maintained at 20/c moisture. Plants were harvested 34 days after